The Genome Sequence of Yaba-Like Disease Virus, a Yatapoxvirus

Home , Yaba monkey tumor virus, Yatapoxvirus

Virology 281, 170–192 (2001) doi:10.1006/viro.2000.0761, available online at http://www.idealibrary.com on

View metadata, citation and similar papers at core.ac.uk brought to you by CORE

provided by Elsevier - Publisher Connector The Genome Sequence of Yaba-like Disease Virus, a Yatapoxvirus

Han-Joo Lee,*,1 Karim Essani,† and Geoffrey L. Smith*,2

*Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, United Kingdom; and †Department of Biological Sciences, Western Michigan University, Michigan 49008 Received July 18, 2000; returned to author for revision August 18, 2000; accepted November 28, 2000

The genome sequence of Yaba-like disease virus (YLDV), an unclassified member of the yatapoxvirus genus, has been determined. Excluding the terminal hairpin loops, the YLDV genome is 144,575 bp in length and contains inverted terminal repeats (ITRs) of 1883 bp. Within 20 nucleotides of the termini, there is a sequence that is conserved in other poxviruses and is required for the resolution of concatemeric replicative DNA intermediates. The nucleotide composition of the genome is 73% AϩT, but the ITRs are only 63% AϩT. The genome contains 151 tightly packed open reading frames (ORFs) that either are Ն180 nucleotides in length or are conserved in other poxviruses. ORFs within 23 kb of each end are transcribed toward the termini, whereas ORFs within the central region of the genome are encoded on either DNA strand. In the central region ORFs have a conserved position, orientation, and sequence compared with vaccinia virus ORFs and encode many enzymes, transcription factors, or structural proteins. In contrast, ORFs near the termini are more divergent and in seven cases are without counterparts in other poxviruses. The YLDV genome encodes several predicted immunomodulators; examples include two proteins with similarity to CC chemokine receptors and predicted secreted proteins with similarity to MHC class I antigen, OX-2, interleukin-10/mda-7, poxvirus growth factor, serpins, and a type I interferon-binding protein. Phylogenic analyses indicated that YLDV is very closely related to yaba monkey tumor virus, but outside the yatapoxvirus genus YLDV is more closely related to swinepox virus and leporipoxviruses than to other chordopoxvirus genera. © 2001 Academic Press Key Words: yatapoxvirus; genome sequence; phylogeny; virus evolution; immune evasion.

INTRODUCTION 1994), India-1968 (Shchelkunov et al., 1995), and Garcia- 1966 (Shchelkunov et al., 2000), molluscum contagiosum Poxviruses are large DNA viruses that infect insects virus (MCV) (Senkevich et al., 1997), Melanoplus sangui- (entomopoxviruses) or vertebrates (chordopoxviruses) nipes entomopoxvirus (MSV) (Afonso et al., 1999), Shope (Fenner, 1996). The chordopoxvirus group is divided into fibroma virus (SFV) (Willer et al., 1999), myxoma virus eight genera, orthopox, parapox, capripox, leporipox, sui- (MYX) (Cameron et al., 1999), fowlpox virus (FPV) (Afonso pox, avipox, molluscipox, and yatapox, of which the or- et al., 2000), and camelpox virus (CMPV) (Gubser and thopoxviruses have been the most important for humans and include variola virus (VAR) (the cause of smallpox), Smith, submitted for publication). vaccinia virus (VV) (the vaccine used to eradicate small- The yatapoxvirus genus contains three members: ta- pox), cowpox virus (CPV), and monkeypox virus. All pox- napox virus (TPV), yaba monkey tumor virus (YMTV), and viruses replicate in the cell cytoplasm and encode their yaba-like disease virus (YLDV) (Knight et al., 1989). All own enzymes for transcription and DNA replication these viruses are morphologically similar to orthopoxvi- (Moss, 1996). ruses but are antigenically distinct from members of this The nucleotide sequence of several poxviruses has and other poxvirus genera and hence cannot be neutral- been determined. These include VV strains Copenhagen ized by polyclonal antisera raised against VV, MCV, or orf (CPN) (Goebel et al., 1990), modified virus Ankara virus (OV), a parapoxvirus (Downie et al., 1971). Yatapox- (Antoine et al., 1998), and the majority of Western Re- viruses have a narrow host range and infect monkeys but serve (WR) (Smith et al., 1991, and references therein), can also be transmitted to humans where they may VAR strains Bangladesh-1975 (VAR-BSH) (Massung et al., cause a mild febrile illness lasting a few days with one or two lesions (deHarven and Yohn, 1966; Espan˜a et al., 1971). The progression of yatapoxvirus-induced pathol- Sequence data from this article have been deposited with the EMBL ogy is considerably slower than that for orthopoxviruses. Data Library under Accession No. AJ293568. TPV was isolated from the skin lesions of patients during 1 Present address: Memorial Slone-Kettering Cancer Center, 1275 outbreaks in the Tana River valley in Kenya in 1957 to York Avenue, RRL 629, Box 362, New York, NY 10021. 1962 (Downie et al., 1971). YMTV was isolated from a 2 To whom correspondence and reprint requests should be ad- dressed at present address: Wright-Fleming Institute, Imperial College colony of rhesus monkeys near Lagos, West Africa in School of Medicine, St. Mary’s Campus, Norfolk Place, London W2 1958 (Bearcroft and Jamieson, 1958) and can produce 1PG, United Kingdom. Fax: 0207-592-3973. E-mail: [email protected]. tumor-like lesions with masses of mesodermal tissue

(Niven et al., 1961). YLDV was isolated in 1965–1966 from monkeys and animal handlers in primate centers in the United States (Casey et al., 1967; Hall and McNulty, 1967; Nicholas and McNulty, 1968; Espan˜a, 1971). The YLDV- induced lesions in monkeys and clinical signs in humans are similar to TPV and these viruses are biologically indistinguishable (McNulty et al., 1968; Downie and Es- pan˜a, 1972). Serological cross-reactivity between yatapoxviruses has been reported (Nicholas and McNulty, 1968; Kupper et al., 1970). Yatapoxviruses have a 145-kb, linear, double-stranded (ds) DNA genome with covalently closed hairpin ends and variations have been noted in the restriction patterns of the different viruses (Knight et al., 1989). Cross-hybridization experiments indicated some sequence conservation between YMTV and monkeypox virus (Rouhandeh et al., 1982), but until recently very little sequence data were available for the yatapoxviruses. Part of the YMTV genome was deposited in a public database (Amano et al., 1995, 2000a, b; Amano and Miyamura, 2000) and small fragments of the TPV genome have been sequenced (Neering and Essani, 1999). To gain an increased understanding of the yatapoxviruses and their evolutionary relationships with other poxviruses, we have determined the sequence of YLDV with FIG. 1. Schematic representation of the YLDV ITRs. Each ITR is the exception of the terminal hairpins. The genome is shown together with the positions of the terminal ORFs, the concate- 144,575 bp and contains 151 open reading frames mer resolution sequence, and the presumed terminal hairpin. Shown (ORFs). The ORFs within the central 100 kb of the ge- beneath the right ITR is the concatemeric resolution sequence from VV, nome show considerable similarity to corresponding MCV, FPV, SFV, and YLDV. Nucleotides conserved in at least four of five sequences are boxed. ORFs in VV in their deduced amino acid sequence, position, and direction of transcription. Several potential immunomodulators are identified and the evolutionary of the virus genome is unknown. However, several lines relationship of YLDV with other poxviruses is discussed. of evidence indicate that the sequence obtained came very close to the terminal hairpins. First, the length ob- RESULTS AND DISCUSSION tained (144,575 bp) was very similar to the total length of the YLDV genome (146.5 bp) estimated from restriction Genome structure of YLDV enzyme digestion and gel electrophoresis (Knight et al., A total of 1,098,770 nucleotides of YLDV DNA were 1989). Second, the size of the terminal PstI restriction assembled into a contiguous sequence of 144,575 bp. fragments described by others (Knight et al., 1989) and Ninety-seven point nine percent of these data were ob- observed experimentally by us was indistinguishable tained from sequencing random clones and the remain- from the length predicted by computer from the se- ing 2.1% were obtained from direct sequencing of either quence generated here. Third, and most compelling, a genomic DNA or 10-kb clones using specific oligonucle- sequence motif was identified within 20 nucleotides of otide primers. The average character density was 7.6 per each terminus that is conserved perfectly with se- nucleotide and 99.0% of the sequence was obtained from quences found very close to the terminal hairpins of both DNA strands. Every position was sequenced at other poxviruses and known to be required for the res- least twice. The genome was composed of 73% AϩT, olution of concatemeric DNA replicative intermediates making YLDV the most AϩT-rich chordopoxvirus to have (Merchlinsky and Moss, 1989) (Fig. 1). In VV strains WR been sequenced. Only the genome of the entomopoxvi- (Baroudy et al., 1982) and CPN (Goebel et al., 1990) and rus MSV has a higher AϩT content (81%) (Afonso et al., VAR-BSH (Massung et al., 1994) this conserved se- 1999). quence is located within 60 bp of the terminal hairpins. The sequence obtained did not include the terminal The motif is also present in FPV (Afonso et al., 2000), SFV hairpins because, as expected, these were not cloned and MYX (Cameron et al., 1999; Willer et al., 1999), and into either library and attempts to sequence these di- MCV (Senkevich et al., 1996). This observation extended rectly using virus DNA as template and specific oligonu- the number of chordopoxvirus genera known to contain cleotides were unsuccessful. Therefore, the exact length this sequence and suggested that the mechanism of 172 LEE, ESSANI, AND SMITH

FIG. 2. YLDV genetic map. Each ORF is represented by an arrow that indicates the approximate size and direction of transcription. Colors indicate proteins with similar functions. ITRs are shown as black bars at the ends of the genome. Each full-length line shows 20 kb.

YLDV DNA replication shares common features with were proposed to contribute to genome length hetero- other poxviruses. geneity. Repeated sequences are also present within the The YLDV genome contains inverted terminal repeats MSV ITR (Afonso et al., 1999). The YLDV ITR is predicted (ITRs) of 1883 bp (Fig. 1). This compares with ITRs of 725 to encode protein to within 758 nucleotides of the termi- bp in VAR-BSH (Massung et al., 1994), 12 kb in VV strain nus of the determined sequence, whereas some other CPN (Goebel et al., 1990), and 50 kb in some mutants of poxviruses, such as VV, have longer noncoding regions CPV (Pickup et al., 1984). The YLDV ITR lacks repeated in the ITR (Goebel et al., 1990). A notable feature of the sequences such as the multiple copies of 50 to 70 YLDV genome was that the ITR had a lower AϩT content nucleotides in the ITRs of several orthopoxviruses (Ba- (63%) than the genome as a whole (73%). The signifi- roudy and Moss, 1982; Goebel et al., 1990; Massung et cance of this is unknown. Similarly, the leporipoxviruses al., 1994). In these viruses these repeated sequences MYX (Cameron et al., 1999) and SFV (Willer et al., 1999) GENOME SEQUENCE OF YLDV 173

TABLE 1

Properties of YLDV ORFs and the Encoded Proteins

ORFa Start/stop AAb kDac IPd TOE Features, putative function Identity/similarity Reference

1L 1,756–755 333 38.5 5.5 E VV C10L (330 aa) 27%/49% in 333 Goebel et al. (1990) 2L 2,885–1,869 338 38.8 5.4 ? Soluble MHC-I protein (?), 1 Ig domain, SP SPV C1L/murine IgE C chain (340 aa) 38%/53% in 336 Massung et al. (1993) Chicken MHC-I glycoprotein (355 aa) 22%/44% in 341 Hunt and Fulton (1998) 3L 3,600–2,932 222 26.6 9.1 L SPV C5L (236 aa) 43%/65% in 215 Massung et al. (1993) VV K7R (149 aa) 24%/53% in 149 Goebel et al. (1990) 4L 4,346–3,630 238 28.5 6.3 L VV N2L (175 aa)/␣-amanitin sensitivity 25%/60% in 174 Tamin et al. (1991) SPV C5L (236 aa) 32%/55% in 222 Massung et al. (1993) 5L 4,856–4,386 156 18.3 8.0 L 2 TMs SPV C7L (155 aa) 32%/53% in 155 Massung et al. (1993) 6L 5,369–4,920 149 17.2 6.0 E VV B15R (149 aa) 40%/61% in 139 Goebel et al. (1990) CPV B13R (149 aa) 41%/62% in 139 Shchelkunov et al. (1998) 7L 6,488–5,433 351 41 9.5 L CC chemokine receptor, 7 TMs Macaca mulatta chemokine receptor 57%/77% in 309 Margulies et al. (1999) Human CC chemokine receptor type 8 (355 aa) 53%/73% in 343 Tiffany et al. (1997) CAPV Q2/3L/G-protein-coupled receptor (381 aa) 39%/65% in 229 Cao et al. (1995) SPV K2R/G-protein-coupled receptor (370 aa) 35%/61% in 368 Massung et al. (1993) 8L 7,146–6,505 213 25.1 5.2 E 4 ankyrin domains CAPV Q2/2L (202 aa) 40%/61% in 197 Cao et al. (1995) VV M1L (421 aa, WR)/ankyrin repeat region 28%/53% in 260 Tamin et al. (1988) 9L 7,863–7,183 226 25.6 7.8 ? SP, VV M2L (220 aa) 28%/50% in 219 Goebel et al. (1990) 10L 9,036–7,885 383 43.6 10.3 L SERP-I, serpin domain, SP MYX SERP-1 (369 aa) 33%/55% in 355 Upton et al. (1990a) VV K2L/SPI-3 (369 aa) 29%/52% in 367 Boursnell et al. (1988) 11L 10,961–9,048 637 73.4 9.9 ? 14 ankyrin domains Human ankyrin (1856 aa) 25%/49% in 726 Forget et al. (1997) VV B4R (558 aa) 25%/48% in 552 Howard et al. (1991) 12L 11,247–10,981 88 9.8 9.8 E eIF2␣ mimic/resistance to IFN SPV C8L (86 aa) 44%/66% in 82 Massung et al. (1993) VV K3L (88 aa)/eIF2␣ 37%/55% in 81 Beattie et al. (1991) 13L 12,136–11,279 285 32.7 9.8 L Monoglyceride lipase, zinc finger motif Mouse monoglyceride lipase (303 aa) 37%/60% in 278 Karlsson et al. (1997) CPV M5L (276 aa) 38%/51% in 276 Shchelkunov et al. (1998) 14L 12,569–12,159 136 16.1 9.9 ? IL-18-binding protein, SP SPV C9L (134 aa) 33%/50% in 127 Massung et al. (1993) MCV MC053L (133 aa) 27%/38% in 92 Senkevich et al. (1997) MCV MC054L (235 aa) 24%/34% in 122 Senkevich et al. (1997) Human IL-18-binding protein (192 aa) 24%/33% in 136 Novick et al. (1999) Mouse IL-18-binding protein (193 aa) 21%/33% in 136 Novick et al. (1999) 15L 12,830–12,597 77 8.8 8.6 ? Growth factor, EGF domain, SP MYX growth factor (85 aa) 40%/60% in 80 Upton et al. (1987) 16L 13,360–12,830 176 20.5 7.7 L Virulence factor, 2 TMs SPV C10L (167 aa) 28%/59% in 166 Massung et al. (1993) MYX M11L (166 aa)/virulence factor 25%/49% in 161 Graham et al. (1992) 17L 13,836–13,405 143 15.4 8.4 ? dUTPase SPV dUTPase (142 aa) 64%/77% in 142 Massung et al. (1993) VV F2L (147 aa)/dUTPase 56%/74% in 140 Broyles (1993) 18L 14,261–13,878 127 15 10.6 ? SPV C12L (75 aa) 32%/54% in 75 Massung et al. (1993) 19L 15,861–14,293 522 60.2 7.1 ? Kelch-like protein SPV C13L (500 aa) 38%/61% in 494 Massung et al. (1993) VV C2L (512 aa) 25%/48% in 484 Goebel et al. (1990) VV A55R (564 aa) 24%/46% in 527 Goebel et al. (1990) 20L 16,863–15,886 325 37.8 5.3 L Ribonucleotide reductase small subunit VV F4L (319 aa)/ribonucleotide reductase small 77%/85% in 317 Roseman and Slabaugh (1990) subunit 21L 17,139–16,891 82 9.4 7.5 ? TM SPV C15L (86 aa) 34%/68% in 86 Massung et al. (1993) 22L 17,495–17,184 103 12.2 5.9 ? 23L 17,712–17,497 71 8.5 10.0 ? 24L 18,732–18,088 214 24.3 8.3 ? TM VV F9L (212 aa) 52%/76% in 212 Goebel et al. (1990) SPV C19L (215 aa) 49%/74% in 215 Massung et al. (1993) 25L 20,047–18,710 445 52.9 7.0 L Serine/threonine protein kinase SPV C20L (440 aa)/possible protein kinase 76%/87% in 440 Massung et al. (1993) VV F10L (439 aa)/serine/threonine kinase 2 72%/83% in 439 Lin and Broyles (1994) 26L 21,999–20,071 642 73.5 8.9 ? 4 TMs VV F12L (635 aa) 38%/59% in 630 Goebel et al. (1990) 27L 23,143–22,031 370 41.5 7.2 L EEV protein MYX MF13 (371 aa)/envelope protein 72%/83% in 367 Jackson and Hall (1998) VV F13L (372 aa)/37-kDa EEV antigen 58%/74% in 367 Hirt et al. (1986) 28R 23,215–23,436 73 8.8 10.6 ? TM 29L 24,036–23,590 148 17.2 9.7 E TM VV F15L (158 aa) 57%/74% in 146 Goebel et al. (1990) 30L 24,748–24,101 215 24.8 10.7 ? SP VV F16L (231 aa) 41%/60% in 224 Goebel et al. (1990) 174 LEE, ESSANI, AND SMITH

TABLE 1—Continued

ORFa Start/stop AAb kDac IPd TOE Features, putative function Identity/similarity Reference

31R 24,807–25,124 105 11.8 9.9 L DNA-binding phosphoprotein MYX MF17 (102 aa) 72%/80% in 100 Jackson and Hall (1998) VV F17R (101 aa)/DNA-binding 11-kDa phosphoprotein 55%/72% in 101 Kao and Bauer (1987) (VP11) 32L 26,533–25,121 470 54.2 9.7 L Poly(A) polymerase catalytic subunit VV E1L (479 aa)/poly(A) polymerase catalytic subunit 66%/81% in 468 Gershon et al. (1991) (VP55) 33L 28,607–26,550 685 79.8 7.8 L VV E2L (737 aa) 42%/66% in 730 Goebel et al. (1990) 34L 29,224–28,667 185 21.3 9.5 ? dsRNA-binding protein/resistance to IFN VV E3L (190 aa)/dsRNA-binding protein/resistance to 38%/55% in 187 Chang et al. (1992, 1995) IFN/host range OV OV20.0L (183 aa)/resistance to IFN 28%/50% in 183 Haig et al. (1998) 35L 29,833–29,264 189 22 9.2 ? RNA polymerase subunit rpo30/intermediate transcription factor-1 VV E4L (259 aa)/RNA polymerase 30-kDa 71%/81% in 186 Ahn et al. (1990a); Rosales et al. subunit/intermediate transcription factor-1 (1994) 36R 29,950–30,996 348 41.2 10.2 ? CMPV E5 (329 aa) 26%/50% in 322 Douglas and Dumbell (1996) VV E5R (331 aa) 24%/47% in 324 Goebel et al. (1990) 37R 30,997–32,700 567 67.1 7.1 L VV E6R (567 aa) 62%/81% in 567 Goebel et al. (1990) 38R 32,720–33,526 268 31.5 8.9 ? Homeobox domain signature VV E8R (273 aa) 69%/80% in 263 Goebel et al. (1990) 39L 36,543–33,523 1006 116.9 8.3 ? DNA polymerase VV E9L (1006 aa)/DNA polymerase 69%/81% in 1005 Earl et al. (1986) FPV DNA polymerase (988 aa) 51%/68% in 1006 Binns et al. (1987) 40R 36,577–36,861 94 10.9 8.8 ? TM, SP VV E10R (95 aa) 66%/78% in 95 Goebel et al. (1990) 41L 37,247–36,858 129 15.2 5.3 L TM VV E11L (129 aa) 43%/70% in 129 Goebel et al. (1990) 42L 39,273–37,234 679 78 10.1 E VV O1L (666 aa) 39%/63% in 666 Goebel et al. (1990) 43L 40,377–39,442 311 36 9.0 L DNA-binding core protein VV I1L (312 aa)/DNA-binding core protein 69%/83% in 311 Klemperer et al. (1997) 44L 40,605–40,378 75 8.5 4.5 L TM VV I2L (73 aa) 47%/74% in 72 Goebel et al. (1990) 45L 41,406–40,606 266 29.7 6.6 E ssDNA-binding phosphoprotein VV I3L (269 aa)/ssDNA-binding phosphoprotein 54%/71% in 269 Davis and Mathews (1993); Rochester and Traktman (1998) 46L 41,710–41,471 79 8.5 10.6 L Structural protein, TM, SP VV I5L (79 aa)/VP13 43%/72% in 75 Takahashi et al. (1994) 47L 42,888–41,728 386 44.9 8.8 L VV I6L (382 aa) 53%/74% in 386 Schmitt and Stunnenberg (1988) 48L 44,174–42,885 429 50.1 8.4 L Topoisomerase II VV I7L (423 aa)/topoisomerase II 69%/85% in 429 Kane and Shuman (1993) 49R 44,180–46,213 677 78.6 10.1 L RNA and DNA helicase VV I8R (676 aa)/RNA & DNA helicase 58%/74% in 672 Shuman (1992); Bayliss and Smith (1996) 50L 47,982–46,210 590 68.6 8.2 L Protease VV G1L (591 aa)/protease 55%/76% in 591 Whitehead and Hruby (1994) 51L 48,314–47,979 111 13 9.6 L SP VV G3L (111 aa) 45%/63% in 111 Goebel et al. (1990) 52R 48,308–48,976 222 25.5 10.3 I Late transcription elongation factor, SP VV G2R (220 aa)/late transcription elongation factor 49%/72% in 220 Condit et al. (1996) 53L 49,320–48,943 125 14.5 5.1 L Glutaredoxin 2 VV G4L (124 aa)/glutaredoxin 2 43%/65% in 124 Gvakharia et al. (1996) 54R 49,323–50,642 439 51.1 8.7 ? VV G5R (434 aa) 45%/65% in 433 Goebel et al. (1990) 55R 50,645–50,836 63 7.3 8.2 ? RNA polymerase subunit rpo7 VV G5.5R (63 aa)/RNA polymerase 7-kDa subunit 79%/88% in 63 Amegadzie et al. (1992) 56R 50,836–51,378 180 20.5 8.9 ? TM VV G6R (165 aa) 49%/71% in 159 Goebel et al. (1990) 57L 52,471–51,347 374 42.6 8.4 L IMV structural protein VV G7L (371 aa)/structural protein, VP16 54%/72% in 367 Takahashi et al. (1994) 58R 52,501–53,283 260 29.8 8.0 I/L Late transcription factor VV G8R (260 aa)/VLTF-1 83%/94% in 260 Keck et al. (1990) 59R 53,296–54,300 334 38.4 7.1 L Myristylprotein, TM VV G9R (340 aa)/myristylprotein 51%/69% in 340 Martin et al. (1997) 60R 54,301–55,044 247 26.8 6.5 L Myristylated IMV protein, TM VV L1R (250 aa)/25-kDa myristylprotein 67%/80% in 247 Franke et al. (1990) 61R 55,063–55,338 91 11 8.4 L 2 TMs VV L2R (87 aa) 28%/70% in 85 Goebel et al. (1990) 62L 56,264–55,314 316 36.9 8.3 L VV L3L (350 aa) 51%/72% in 319 Goebel et al. (1990) 63R 56,289–57,038 249 28.6 5.9 L IMV structural protein VV L4R (251 aa)/IMV structural protein VP8, DNA/RNA- 60%/82% in 247 Yang and Bauer (1988); Bayliss binding protein and Smith (1997) 64R 57,054–57,440 128 14.8 9.7 L TM VV L5R (128 aa) 46%/68% in 128 Goebel et al. (1990) GENOME SEQUENCE OF YLDV 175

TABLE 1—Continued

ORFa Start/stop AAb kDac IPd TOE Features, putative function Identity/similarity Reference

65R 57,394–57,873 159 18.6 5.4 ? CAPV F7 (147 aa) 67%/77% in 147 Gershon and Black (1989) MYX MF7 (148 aa) 59%/70% in 148 Jackson and Bults (1992) VV J1R (153 aa) 49%/66% in 153 Goebel et al. (1990) 66R 57,870–58,421 183 20.5 8.4 ? Thymidine kinase YMTV thymidine kinase (181 aa) 82%/90% in 183 Amano et al. (1995) VV J2R (177 aa)/thymidine kinase 68%/77% in 175 Hruby et al. (1983); Weir and Moss (1983) 67R 58,469–59,005 178 21 4.5 E Host range protein SPV SWF8A (185 aa) 45%/68% in 179 Schnitzlein and Tripathy (1991) VV C7L (150 aa)/host range protein 35%/66% in 150 Perkus et al. (1990) 68R 58,983–60,074 363 42.8 10.4 ? Poly(A) polymerase regulatory subunit MYX F9 (338 aa) 73%/85% in 333 Jackson and Bults (1990) VV J3R (333 aa)/poly(A) polymerase regulatory subunit 71%/85% in 333 Gershon and Moss (1993) (VP39) 69R 59,989–60,546 185 21.1 6.8 ? RNA polymerase subunit rpo22 VV J4R (185 aa)/RNA polymerase 22-kDa subunit 74%/88% in 185 Broyles and Moss (1986) 70L 60,936–60,523 137 15.9 7.8 L 1 TM VV J5L (133 aa) 62%/78% in 130 Goebel et al. (1990) 71R 61,027–64,884 1285 147.6 8.8 E RNA polymerase subunit rpo147 VV J6R (1286 aa)/RNA polymerase 147-kDa subunit 80%/89% in 1286 Broyles and Moss (1986) RNA polymerase subunit family Ͻ80% 72L 65,396–64,881 171 19.9 10.1 L Tyrosine/serine phosphatase VV H1L (171 aa)/tyrosine/serine phosphatase 64%/83% in 171 Guan et al. (1991) 73R 65,412–65,984 190 21.9 9.3 ? TM VV H2R (189 aa) 62%/78% in 188 Goebel et al. (1990) 74L 66,958–65,987 323 37.3 6.8 L Immunodominant envelope protein, TM VV H3L (324 aa)/immunodominant envelope protein 35%/64% in 320 Chertov et al. (1991) p35 75L 69,355–66,959 798 94.7 7.7 L RNA polymerase-associated transcription specificity factor (RAP) 94 VV H4L (795 aa)/RAP 94 69%/84% in 798 Ahn and Moss (1992); Kane and Shuman (1992) 76R 69,514–70,056 180 20.2 9.0 I Late transcription factor VLTF-4 VV H5R (203 aa)/late transcription factor-4 46%/61% in 203 Kovacs and Moss (1996) 77R 70,069–71,016 315 36.9 10.5 I/L DNA topoisomerase I SFV DNA topoisomerase I (314 aa) 64%/81% in 314 Upton et al. (1990b) VV H6R (314 aa)/DNA topoisomerase I 63%/77% in 313 Shuman and Moss (1987) 78R 71,032–71,478 148 17 9.9 L VV H7R (146 aa) 39%/62% in 143 Goebel et al. (1990) 79R 71,516–74,038 840 97.9 9.0 L mRNA capping enzyme large subunit SFV mRNA capping enzyme large subunit (836 aa) 69%/84% in 834 Upton et al. (1991) VV D1R (844 aa)/mRNA capping enzyme large subunit 66%/81% in 844 Morgan et al. (1984) 80L 74,461–74,000 154 18.1 10.1 L Structural protein VV D2L (146 aa)/structural protein 52%/73% in 139 Dyster and Niles (1991) 81R 74,460–75,197 245 28.4 7.5 ? Structural protein VV D3R (237 aa)/structural protein 39%/58% in 245 Dyster and Niles (1991) 82R 75,194–75,850 218 25.1 8.5 L Uracil-DNA glycosylase SFV uracil-DNA glycosylase (218 aa) 72%/87% in 218 Upton et al. (1993) VV D4R (218 aa)/uracil-DNA glycosylase 69%/88% in 218 Stuart et al. (1993) 83R 75,956–78,316 786 90.1 7.8 ? NTPase, ATP/GTP-binding site motif A (P-loop) VV D5R (785 aa)/NTPase 69%/82% in 785 Evans et al. (1995) 84R 78,313–80,220 635 73.1 9.1 L Early transcription factor SFV early transcription factor 70-kDa subunit (635 aa) 86%/93% in 635 Strayer et al. (1991) VV D6R (637 aa)/early transcription factor 70-kDa 80%/90% in 637 Broyles and Fesler (1990); subunit Gershon and Moss (1990) 85R 80,253–80,735 160 17.9 5.8 L RNA polymerase subunit rpo18 VV D7R (161 aa)/RNA polymerase 18-kDa subunit 71%/84% in 160 Ahn et al. (1990b); Quick and Broyles (1990) 86R 80,780–81,418 212 24.7 8.6 ? mutT protein, mutT domain signature VV D9R (213 aa)/mutT 60%/76% in 206 Koonin (1993) 87R 81,415–82,182 255 29.9 10.2 L Negative regulator of gene expression, mutT domain signature SFV D10R (260 aa)/mutT 60%/75% in 250 Strayer et al. (1991) VV D10R (248 aa)/negative regulator of gene 51%/69% in 248 Shors et al. (1999) expression 88L 84,070–82,175 631 72.8 10.0 L RNA polymerase elongation factor VV D11L (631 aa)/NPH-I 70%/83% in 631 Christen et al. (1998); Deng and Shuman (1998) 89L 84,965–84,102 287 33.4 8.1 I/L mRNA capping enzyme small subunit SPV mRNA capping enzyme small subunit (287 aa) 82%/93% in 287 Massung et al. (1993) VV D12L (287 aa)/mRNA capping enzyme small 74%/89% in 287 Niles et al. (1989) subunit 90L 86,652–84,994 552 62.3 6.5 L Rifampicin resistance SPV rifampicin resistance protein (551 aa) 78%/89% in 550 Massung et al. (1993) VV D13L (551 aa)/rifampicin resistance 73%/84% in 551 Tartaglia and Paoletti (1985) 176 LEE, ESSANI, AND SMITH

TABLE 1—Continued

ORFa Start/stop AAb kDac IPd TOE Features, putative function Identity/similarity Reference

91L 87,128–86,676 150 17 9.0 I/L Late gene transcription factor VV A1L (150 aa)/VLTF-2 64%/85% in 150 Keck et al. (1993) 92L 87,826–87,152 224 26.1 8.6 I/L Late gene transcription factor VV A2L (224 aa)/VLTF-3 84%/95% in 224 Hubbs and Wright (1996) 93L 88,050–87,823 75 9.1 10.2 L VV WR hypothetical 8.9-kDa protein (76 aa) 56%/69% in 76 Weinrich and Hruby (1986) 94L 90,033–88,060 657 75.4 6.9 L Core structural protein VV A3L (644 aa)/core structural protein P4b 65%/81% in 657 Rosel and Moss (1985) 95L 90,548–90,090 152 17.5 5.1 ? Membrane-associated protein VV A4L (281 aa)/membrane-associated protein 30%/65% in 76 Cudmore et al. (1996) 96R 90,588–91,094 168 19.2 4.2 ? RNA polymerase subunit rpo19 VV A5R (164 aa)/RNA polymerase 19-kDa subunit 63%/80% in 168 Ahn et al. (1992) 97L 92,206–91,091 371 43.2 8.8 I/L VV A6L (372 aa) 58%/77% in 368 Goebel et al. (1990) 98L 94,365–92,230 711 81.9 8.5 L Early transcription factor 82-kDa subunit VV A7L (710 aa)/early transcription factor 82-kDa 71%/85% in 711 Gershon and Moss (1990) subunit 99R 94,418–95,290 290 33.9 9.2 L Intermediate gene transcription factor subunit VV A8R (288 aa)/VITF-3 61%/78% in 288 Sanz and Moss (1999) 100L 95,526–95,287 79 9 8.7 L TM, SP VV A9L (99 aa) 73%/86% in 90 Goebel et al. (1990) 101L 98,235–95,527 902 103 6.4 L Core structural protein P4a VV A10L (891 aa)/core structural protein P4a 54%/72% in 902 Van Meir and Wittek (1988) 102R 98,250–99,185 311 34.9 5.6 L VV A11R (318 aa) 57%/79% in 318 Goebel et al. (1990) 103L 99,707–99,195 170 18.3 10.4 ? Structural protein VV A12L (192 aa)/structural protein 54%/70% in 191 Takahashi et al. (1994) 104L 99,956–99,750 68 7.7 9.2 ? IMV membrane protein, TM VV A13L (70 aa)/IMV membrane protein P8 31%/55% in 70 Jensen et al. (1996) 105L 100,285–100,004 93 10.1 8.7 ? IMV membrane protein, 2 TMs VV A14L (90 aa)/IMV membrane protein P16 56%/72% in 90 Jensen et al. (1996) 106L 100,453–100,302 53 6.2 8.4 L IMV membrane protein, SP, TM VV A14.5L (53 aa)/IMV membrane protein 52%/63% in 51 Betakova et al. (2000) 107L 100,737–100,453 94 10.8 9.3 L VV A15L (94 aa) 52%/68% in 94 Goebel et al. (1990) 108L 101,866–100,721 381 44.6 8.8 L Myristylprotein, TM VV A16L (378 aa)/myristylprotein 52%/71% in 381 Martin et al. (1997) 109L 102,455–101,880 191 20.9 4.8 L IMV membrane protein, 4 TMs VV A17L (203 aa)/IMV membrane protein 41%/65% in 197 Rodriguez et al. (1995) 110R 102,470–103,909 479 55.6 10.8 I/L DNA helicase VV A18R (493 aa)/DNA helicase 56%/71% in 493 Simpson and Condit (1995) 111L 104,114–103,890 74 7.9 9.8 ? VV A19L (77 aa) 63%/79% in 69 Goebel et al. (1990) 112L 104,447–104,115 110 12.7 8.6 L SP VV A21L (117 aa) 59%/72% in 117 Goebel et al. (1990) 113R 104,446–105,723 425 49 8.9 ? DNA polymerase processivity factor VV A20R (426 aa)/DNA polymerase processivity factor 47%/67% in 424 McDonald et al. (1997) 114R 105,732–106,205 157 18.5 10.7 L TM VV A22R (176 aa) 67%/85% in 157 Goebel et al. (1990) 115R 106,227–107,378 383 44.5 10.3 L Intermediate gene transcription factor subunit VV A23R (382 aa)/VITF-3 61%/77% in 380 Sanz and Moss (1999) 116R 107,380–110,877 1165 133.8 8.1 L RNA polymerase subunit rpo132 CPV RNA polymerase 132-kDa subunit (1164 aa) 82%/91% in 1164 Patel and Pickup (1989) VV A24R (1164 aa)/RNA polymerase 132-kDa subunit 82%/91% in 1163 Amegadzie et al. (1991) CAPV RNA polymerase subunit (919 aa) 87%/93% in 919 Gershon et al. (1989) 117L 111,320–110,874 148 17.2 5.0 ? IMV surface protein CAPV HM2 (148 aa)/putative fusion protein 42%/62% in 146 Gershon et al. (1989) VV A27L (110 aa)/IMV surface protein 39%/55% in 106 Rodriguez and Smith (1990) 118L 111,740–111,321 139 16.1 6.5 L SP MYX MA28L (140 aa)/hypothetical 16-kDa protein 63%/80% in 140 Jackson et al. (1996) CAPV HM3 (140 aa) 62%/81% in 140 Gershon et al. (1989) VV A28L (146 aa) 58%/73% in 146 Goebel et al. (1990) 119L 112,655–112,753 300 35.1 6.2 ? RNA polymerase subunit rpo35 VV A29L (305 aa)/RNA polymerase 35-kDa subunit 57%/75% in 296 Amegadzie et al. (1991) 120L 112,851–112,624 75 8.6 7.5 ? VV A30L (77 aa, WR) 49%/69% in 75 Amegadzie et al. (1991) MCV MC136L (67 aa) 48%/69% in 65 Senkevich et al. (1997) 121L 113,814–113,053 253 29.1 9.7 L ATP/GTP-binding site motif A (P-loop) MCV MC140L (249 aa) 63%/80% in 247 Senkevich et al. (1997) VV A32L (300 aa)/ATP/GTP-binding motif 60%/74% in 254 Koonin et al. (1993); Cassetti et al. (1998) 122R 113,926–114,486 186 21 4.7 L EEV glycoprotein, TM VV A33R (185 aa)/EEV glycoprotein 38%/61% in 178 Roper et al. (1996) 123R 114,509–115,021 170 19.5 10.4 L EEV glycoprotein, 1 TM, C-type lectin domain VV A34R (168 aa)/EEV glycoprotein 50%/68% in 170 Duncan and Smith (1992) 124R 115,059–115,598 179 20.5 6.5 ? VV A35R (176 aa) 40%/61% in 171 Goebel et al. (1990) MCV MC145R (233 aa) 26%/53% in 179 Senkevich et al. (1997) 125R 115,634–116,491 285 32.9 8.3 E MCV MC144R (642 aa) 23%/48% in 285 Senkevich et al. (1997) FPV hypothetical 32.4-kDa protein (283 aa) 26%/48% in 278 Hertig et al. (1997) 126R 116,549–117,220 223 25.9 4.2 ? SP 127R 117,265–118,077 270 31.5 10.1 L VV A37R (263 aa) 25%/53% in 253 Goebel et al. (1990) GENOME SEQUENCE OF YLDV 177

TABLE 1—Continued

ORFa Start/stop AAb kDac IPd TOE Features, putative function Identity/similarity Reference

128L 118,890–118,072 272 32.1 10.4 L CD47, 5 TMs, SP Human CD47 precursor (323 aa) 24%/48% in 311 Campbell et al. (1992) VV A38L (277 aa, WR)/integral membrane glycoprotein 23%/47% in 276 Parkinson et al. (1995) 129R 118,900–119,316 138 16 7.6 L VV E7R (166 aa)/myristylprotein 26%/48% in 145 Martin et al. (1997) 130L 119,862–119,302 186 21.2 9.1 L SP 131R 119,939–120,145 68 7.6 4.7 ? 132R 120,173–120,421 82 9.2 4.8 E 133L 121,458–120,427 343 38.9 7.9 ? 3-␤ hydroxysteroid dehydrogenase VV A44L (346 aa)/3␤-hydroxysteroid dehydrogenase 44%/63% in 343 Moore and Smith (1992) 134R 121,497–121,967 156 18.3 10.0 ? mda-7 or IL-10, SP Human mda-7 protein precursor (206 aa) 27%/55% in 175 Jiang et al. (1996) Human IL-10 precursor (178 aa) 23%/53% in 164 Vieira et al. (1991) 135R 122,043–127,748 1901 217.1 8.2 L 8 TMs, SP MCV MC035R (2133 aa) 32%/53% in 2025 Senkevich et al. (1997) 136R 127,745–128,800 351 41.1 7.6 ? IFN ␣/␤ receptor, 1 Ig domain, SP VV B19R (353 aa)/IFN ␣/␤ receptor 26%/45% in 350 Symons et al. (1995) 137R 128,827–129,288 153 17.6 8.1 E 2 TMs VV C6L (151 aa) 23%/54% in 139 Goebel et al. (1990) 138R 129,314–130,330 338 38 7.1 ? VV A51R (334 aa) 34%/57% in 331 Goebel et al. (1990) 139R 130,405–130,977 190 22.3 7.8 ? VV A52R (190 aa) 31%/59% in 185 Goebel et al. (1990) 140R 131,009–132,721 570 65.2 10.1 ? Kelch-like protein VV A55R (564 aa) 30%/54% in 562 Goebel et al. (1990) SPV C4L (530 aa) 30%/53% in 527 Massung et al. (1993) Kelch protein (689 aa) 24%/48% in 554 Xue and Cooley (1993) 141R 132,748–133,110 120 13.5 10.0 ? Soluble OX-2, 1 Ig domain, SP Human OX-2 membrane glycoprotein precursor 25%/50% in 132 McCaughan et al. (1987) (274 aa) MYX MA56 (218 aa) 41%/46% in 115 Jackson et al. (1999) Rhesus radinovirus R15 (253 aa) 29%/38% in 120 Searles et al. (1999) 142R 133,144–134,073 309 35.9 9.6 ? Serine/threonine protein kinase Human serine/threonine protein kinase VRK1 51%/70% in 306 Nezu et al. (1997) (396 aa) VV B1R (300 aa)/serine/threonine protein kinase 49%/67% in 292 Howard and Smith (1989) 143R 134,106–134,810 234 27.8 9.5 L DNA-binding protein, ring domain SFV N1R (234 aa)/DNA-binding protein 45%/64% in 231 Brick et al. (1998) 144R 134,840–135,643 267 30.8 8.6 L CD46, TM, 3 complement control protein domains, SP Human membrane cofactor protein CD46 (314 aa) 29%/50% in 271 Liszewski et al. (1991) VV B5R (317 aa)/EEV membrane glycoprotein/host 28%/49% in 312 Engelstad et al. (1992) range protein precursor VV WR C21L (263 aa)/complement control protein 24%/44% in 256 Kotwal and Moss (1988) precursor 145R 135,898–136,869 323 37.6 7.5 ? CC chemokine receptor, G-protein-coupled receptors signature, 7 TMs Human CC chemokine receptor type 8 (355 aa) 44%/68% in 325 Tiffany et al. (1997) SPV K2R (370 aa)/G-protein-coupled receptor 36%/69% in 364 Massung et al. (1993) CAPV Q2/3L (381 aa)/G-protein-coupled receptor 35%/63% in 319 Cao et al. (1995) 146R 137,018–138,439 473 55 10.3 ? Virulence factor, 6 ankyrin domains SFV T5 (328 aa)/virulence factor 28%/53% in 326 Mossman et al. (1996) VV C9L (634 aa)/ankyrin-like protein 27%/36% in 417 Goebel et al. (1990) 147R 138,461–139,936 491 57.8 9.3 L Virulence factor, 5 ankyrin domains SFV T5 (328 aa)/virulence factor 25%/53% in 324 Mossman et al. (1996) VV B4R (558 aa)/ankyrin-like protein 23%/50% in 489 Howard et al. (1991) 148R 139,947–141,377 476 55.7 10.5 ? Virulence factor, 6 ankyrin domains SFV T5 (328 aa)/virulence factor 24%/54% in 324 Mossman et al. (1996) VV B4R (558 aa)/ankyrin-like protein 23%/50% in 489 Howard et al. (1991) 149R 141,388–142,392 334 38.1 4.9 E SPI-2, serpin domain MYX putative serpin-like protein (333 aa) 45%/64% in 331 Petit et al. (1996) CPV IL-1␤-converting enzyme (ICE) (caspase 1) 40%/63% in 338 Ray et al. (1992) inhibitor (341 aa) VV WR B13R (345 aa)/SPI-2 41%/62% in 341 Smith et al. (1989) 150R 142,430–142,732 100 12.1 10.5 L TM SPV C2L (92 aa) 41%/69% in 86 Massung et al. (1993) 151R 142,820–143,821 333 38.5 5.5 E VV C10L (330 aa) 27%/49% in 333 Goebel et al. (1990)

Note. TOE, time of expression as predicted as described in text; SP, signal peptide; TM, transmembrane domain; VV, vaccinia virus; CMPV, camelpox virus; SFV, Shope fibroma virus; SPV, swinepox virus; CAPV, capripox virus; MYX, myxoma virus; MCV, molluscum contagiosum virus; YMTV, yaba monkey tumor virus; FPV, fowlpox virus. a ORFs are numbered starting from the left end of the genome and the direction of transcription is indicated by an L or an R. Underlined ORFs are diploid because they are present within the ITR. b The number of amino acids of the YLDV protein. c Molecular mass of each protein calculated by PEPSTATS in the XGCG package. d Predicted isoelectric points of proteins calculated by PEPSTATS. 178 LEE, ESSANI, AND SMITH have ITRs with a lower AϩT content (51.0% and 45.0%) tional start site (TAAAT or TAAAAT) was not found within than the whole genome (56.4 and 60.5%, respectively). the 50 bp upstream of the start codon. Similarly, an ORF The ITRs of FPV are also less AϩT rich than the whole was predicted to be expressed late if it contained the late genome (65.4% versus 69.0%) (Afonso et al., 2000). In the start site (TAAAT or TAAAAT) overlapping the transla- orthopoxviruses (Goebel et al., 1990; Shchelkunov et al., tional start codon or within the 50 bp upstream of it and 1995) and MSV (Afonso et al., 1999), the ITRs and whole without an intervening ATG, and if there was one or more ϩ genome have a similar A T content. In contrast, the ITR T5NT motifs within the ORF. Intermediate genes were of MCV has a higher AϩT content (45%) than the whole predicted by the presence of a TAAA initiator sequence genome (36%) (Senkevich et al., 1996). 11–12 bp downstream of the core sequence AAANAA (Baldick et al., 1992). By these criteria 14 ORFs were Arrangement of YLDV ORFs predicted to be early, 72 ORFs late, 2 ORFs intermediate, and 7 ORFs intermediate/late, and 56 ORFs were unas- The YLDV genome contains 150 ORFs of Ն180 nucle- signed (Table 1). The prediction of only 14 early ORFs is otides. In addition an ORF of 159 nucleotides (106L) is very likely to be an underestimate. In VV approximately included because a very closely related protein is ex- half the genome is transcribed early during infection pressed by VV and has conserved counterparts in sev- (Boone and Moss, 1978) and some genes known to be eral other poxviruses (Betakova et al., 2000). ORFs are expressed early in VV, e.g., DNA polymerase and thymi- numbered from 1 to 151 starting from the left terminus dine kinase, are not predicted to be expressed early in and followed by L (left) or R (right) to indicate the direction of transcription (Fig. 2). ORFs 1L and 151R are within YLDV (using the above criteria), but are very likely to be the ITR and therefore are identical, leaving 150 unique so. The time of expression of each gene awaits experi- ORFs. The ORFs are tightly packed and with the excep- mental determination. tion of the terminal 758 bp there is little noncoding DNA. There are only three other positions in the virus genome Analysis of YLDV ORFs where a noncoding DNA region exceeds 200 nucleo- YLDV protein sequences were compared with the tides: between ORFs 23L and 24L (376 nucleotides), 120L SWALL protein database and analyzed for sequence and 121L (201 nucleotides), and 144R and 145R (255 features by XEGCG and SMART programs. Table 1 sum- nucleotides). marizes the position, length, and predicted transcrip- A striking feature of the YLDV genome is that all genes tional regulation of each YLDV ORF and the size, isoelec- located within 23 kb of either terminus (1L to 27L and tric point, amino acid sequence motifs, and similarity 134R to 151R) are transcribed outward toward the with other proteins of each predicted YLDV protein. The genomic termini, whereas genes located with the central ϳ great majority of YLDV proteins (130/151) have counter- 100 kb of the genome (28R to 133R) are transcribed parts in VV with between 24% (148R) and 84% (92L) from either DNA strand. This arrangement is similar to amino acid identity. ORFs without counterparts in VV are that reported for other poxviruses (Goebel et al., 1990; located near either terminus (2L, 5L, 7L, 14L, 15L, 16L, Massung et al., 1994; Senkevich et al., 1996). It was 18L, and 21L from the left end and 125R, 134R, 135R, postulated that the arrangement of genes in this way 141R, 145R, and 150R from the right end) and some of minimizes the production of dsRNA that might induce these ORFs encode proteins with similarity to proteins production of interferon (IFN) or activate IFN-induced, from other viruses or cells. antiviral, dsRNA-binding proteins such as protein kinase Proteins encoded within the central region of the YLDV R (PKR) or 2Ј5Ј-oligoadenylate synthetase (Smith et al., genome show highest amino acid identity with VV coun- 1998). The ORFs encode proteins that range from 53 terparts and with comparable proteins in other poxvi- (106L) to 1901 (135R) amino acids in length with a mean ruses, whereas genes nearer the termini are more vari- size of 305 amino acids, and 71 are transcribed rightward able (Fig. 3). Figure 3 also shows that there are relatively and 80 leftward. few ORFs within the central region of the YLDV genome that do not have a VV counterpart, which is in contrast to Transcriptional regulatory signals the more variable termini. Many of the proteins encoded

The VV early termination sequence motif (T5NT) (Yuen in the central region are enzymes, transcription factors, and Moss, 1987) and the late transcriptional start site or structural proteins that are essential for VV replication TAAAT(G) or TAAAAT(G) (Bertholet et al., 1986; Rosel et (Moss, 1996) (Fig. 2 and Table 1). With few exceptions, al., 1986; Davison and Moss, 1989) were identified near genes in this region are conserved in position, se- the end or at the beginning of many YLDV ORFs. An ORF quence, and orientation compared with VV (Fig. 4). A was considered likely to be expressed early during in- similar conservation of this central block of genes has fection if a T5NT motif was located within the 35 bp been noted in other poxviruses such as MCV and SFV upstream or 50 bp downstream of the stop codon (but (Senkevich et al., 1997; Willer et al., 1999), indicating that not elsewhere within the ORF), and the late transcrip- this region is likely to represent the minimal block of GENOME SEQUENCE OF YLDV 179

FIG. 3. A comparison of YLDV and VV proteins. The amino acid identity (%) of YLDV proteins with their VV counterparts is illustrated. Where no bar is present, the YLDV protein has no counterpart in VV. genes of an ancestral poxvirus to which genus- and lacks a D8L-like protein, it contains counterparts of the virus-specific genes have been added at either terminus other VV IMV surface proteins that bind to cells; e.g., the during evolution. proteins encoded by ORFs 74L and 117L are counter- Despite the considerable similarity with VV in this parts of VV H3L (Lin et al., 2000) and A27L (Chung et al., region, there are also some interesting differences (Fig. 1998), respectively. ORFs A25L and A26L in VV-CPN 4). First, YLDV lacks genes related to VV F11L, F14L, O2L, represent fragments of the gene encoding the A-type I4L, D8L, A25L, A26L, and A31L. The VV D8L protein is inclusion body protein of CPV and some other poxviruses located on the surface of intracellular mature virus (IMV) and which in VV strain WR encodes a 94-kDa protein. particles, is nonessential for virus replication, and en- This gene has been deleted entirely in YLDV and, con- ables IMV particles to bind to cells via chondroitin sulfate sistent with this, electron microscopy of YLDV-infected (Niles and Seto, 1988; Hsiao et al., 1999). Although YLDV owl monkey kidney (OMK) cells revealed no A-type in-

FIG. 4. Alignment of YLDV and VV ORFs from the central genomic region. Arrows represent the direction of transcription and approximate length of each ORF. Black and gray arrows represent YLDV and VV ORFs, respectively. Underlined VV ORFs are absent from the YLDV genome. YLDV ORF 67R is related to VV ORF C7L, which is located in the left region of the genome. 180 LEE, ESSANI, AND SMITH

TABLE 2 YLDV ORFs Predicted to Encode Proteins Involved in Transcription and Their Counterparts in VV, MCV, and MYX

Function YLDV VV MCV MYX

Proteins involved in early transcription RNA polymerase subunits rpo30 35L E4L MC034L M030L rpo7 55R G5.5R MC061R M050R rpo22 69R J4R MC077R M066R rpo147 71R J6R MC079R M068R rpo18 85R D7R MC097R M082R rpo19 96R A5R MC108R M094R rpo132 116R A24R MC129R M114R rpo35 119L A29L MC135L M117L Poly(A) polymerase Catalytic subunit 32L E1L MC031L M027L Regulatory subunit 68R J3R MC076R M065R mRNA capping enzyme Large subunit 79R D1R MC090R M076R Small subunit 89L D12L MC101L M087L RNA polymerase accessory RAP 94 75L H4L MC085L M072L proteins RNA/DNA helicase 49R I8R MC050L M044R Elongation factor 88L D11L MC100L M086L Early transcription factors 84R D6R MC095R M081R 98L A7L MC110L M096L Proteins involved in intermediate Intermediate transcription factor-1 35L E4L MC034L M030L transcription Intermediate transcription factor-3 99R A8R MC111R M097R subunits 115R A23R MC128R M113R Proteins involved in late transcription Late transcription factor-1 58R G8R MC067R M053R Late transcription factor-4 76R H5R MC086R M073R Late transcription factor-2 91L A1L MC103L M089L Late transcription factor-3 92L A2L MC104L M090L Late elongation factor 52R G2R MC058R M047R

clusion bodies (data not shown). VV ORF I4L encodes the their VV counterparts and some of these are considered large subunit of ribonucleotide reductase, a protein that under appropriate sections below. is not encoded in MCV and SFV in addition to YLDV Overall, the predicted YLDV proteins show greatest (Senkevich et al., 1997; Willer et al., 1999). This gene is similarity to proteins from chordopoxviruses and al- nonessential for VV replication in cell culture, although a though in some cases there are related proteins in Afri- VV strain lacking this gene shows a modest attenuation can swine fever virus (ASFV) and MSV, in no case was a (Child et al., 1990). VV ORF O2L encodes glutaredoxin I, YLDV protein more closely related to proteins from these and a related gene is absent in YLDV, MCV (Senkevich et viruses than a chordopoxvirus protein. No YLDV protein al., 1997), MYX (Cameron et al., 1999), and SFV (Willer et was found to be related only to a protein from ASFV or al., 1999). The functions of the other VV genes from the MSV and not from chordopoxviruses. central region of the genome for which YLDV has no counterparts (F11L, F14L, and A31R) are unknown. Transcriptional proteins The central region of the YLDV genome contains a few genes that are transposed or oriented inversely with Sequence comparisons showed that YLDV and VV respect to their VV counterparts (Fig. 4). For instance, the have very similar transcriptional enzymes and associ- VV gene C7L, encoding a host range protein (Perkus et ated factors. These include the eight subunits of RNA al., 1990; Oguiura et al., 1993), is present in the left polymerase (35L, 55R, 69R, 71R, 85R, 96R, 116R, and variable region of the VV genome, but in YLDV this gene 119R), early (84R and 98L), intermediate (35L, 99R, and (67R) is present in the central region immediately to the 115R), and late (52R, 58R, 76R, 91L, and 92L) transcription right of the thymidine kinase gene and is transcribed in factors, RNA polymerase-associated protein (RAP) 94 the opposite direction. The YLDV equivalent of VV E7L is (75L), and the heterodimeric capping (79R and 89L) and also transposed from its position in the VV central region polyadenylating (32L and 68R) enzymes (Table 2). Com- near the right end of the YLDV genome (ORF 129R). In pared with other proteins, these transcriptional enzymes SFV and MYX, genes related to VV C7L and E7L are show very high conservation between VV and YLDV. For present in the same position as YLDV, consistent with instance, the two largest RNA polymerase subunits 71R the closer relationship of YLDV with leporipoxviruses (rpo147) and 115R (rpo132) share more than 80% amino than orthopoxviruses (see Phylogeny below). Several acid identity with their corresponding VV proteins. Most ORFs in the central region are truncated compared to of these proteins are of a very similar length in YLDV and GENOME SEQUENCE OF YLDV 181

VV, a notable exception being 35L (VV rpo30), which is 60 YLDV has a protein (144R) related to complement control amino acids shorter in YLDV. In MCV this protein is proteins (see below), this is more closely related to the longer than the VV counterpart (Senkevich et al., 1997). human protein CD46 than to VV B5R. Thus it is uncertain YLDV transcription factors are also of a similar length to whether the YLDV protein 144R is a functional counter- their VV counterparts, with the exception of the YLDV 76R part of the VV B5R protein, and this awaits experimental protein, which is 20 amino acids shorter than the VV determination. The absence of a counterpart of A36R protein H5R. Overall, these data indicate that YLDV has a protein is interesting. This protein was thought originally transcription system very similar to that of VV. to be part of the EEV envelope because it copurified with EEV in density gradients (Parkinson and Smith, 1994), but Nucleic acid modifying enzymes recently immunogold electron microscopy showed that YLDV, like other poxviruses, has enzymes that are the protein was present on intracellular enveloped virus involved in biosynthesis of nucleic acid precursors. particles but not cell-associated enveloped (CEV) or EEV These include thymidine kinase (66R), the small subunit particles (van Eijl et al., 2000). Instead, it is located on the of ribonucleotide reductase (20L), and dUTPase (17L). cytosolic face of the plasma membrane beneath CEV YLDV lacks counterparts of VV thymidylate kinase and particles. A36R is essential for the formation of virus- the large subunit of ribonucleotide reductase. The latter induced actin tails that drive CEV particles away from the protein is also absent in MCV and SFV (Senkevich et al., cell and are important for efficient cell-to-cell spread of 1997; Willer et al., 1999). The dUTPase gene is nones- virus (Sanderson et al., 1998; Wolffe et al., 1998; Ro¨ttger sential for VV replication but minimizes incorporation of et al., 1999). The absence of an A36R-like protein might UTP into DNA. YLDV also lacks a DNA ligase that is explain why the plaques formed by YLDV take such a found in VV and some other poxviruses, but which is long time to develop, similar to VV plaques formed by nonessential for replication. mutants unable to make actin tails.

DNA replication and repair Host range proteins YLDV encodes proteins related to VV DNA polymerase Several poxvirus host range proteins have been dis- (39L), DNA topoisomerase I (77R), DNA polymerase pro- covered, such as VV K1L and C7L, which are required for cessivity factor (113R), an ATP–GTP-binding protein growth in primate and human cells (Perkus et al., 1990). (83R), and a serine/threonine protein kinase (142R) that YLDV has a protein with 37% amino acid identity to VV are all essential for VV DNA replication. These proteins C7L (67R), but as noted above, this is present in the contained appropriate signatures for their respective central region of the YLDV genome, whereas in VV it is protein families. In addition, YLDV has proteins related to near the left end of the genome. YLDV lacks a counter- uracil DNA glycosylase (82R) and mutT (86R and 87R). part of K1L. Some poxvirus host range proteins contain Uracil DNA glycosylase functions in DNA repair by indi- ankyrin repeats, including K1L and the CPV 77-kDa host rectly causing excision of uracil from DNA, and the mutT range protein (Spehner et al., 1988), and there are sev- proteins are putative NTP pyrophosphohydrolases in- eral other poxvirus proteins of unknown function that volved in preventing DNA damage (Koonin, 1993). An- contain ankyrin repeats, such as VV M1L and B4R other VV protein implicated in repair of damaged DNA, (Goebel et al., 1990). YLDV also has several proteins with DNA ligase (Kerr et al., 1991), is absent from YLDV as ankyrin repeats: Protein 8L has 4 ankyrin repeats and noted above. shares 27% amino acid identity with M1L, and 11L has 14 ankyrin repeats and is more similar to VV B4R. At the Proteins of extracellular enveloped virus right end of the YLDV genome there is a family of related TPV and YLDV make two forms of virus particle that genes (146R, 147R, and 148R) encoding proteins with 5 or have different buoyant densities in CsCl (Knight et al., 6 ankyrin repeats. These share 24 to 29% amino acid 1989) (data not shown). IMV remains in the cell and has identity with one another and so are unlikely to have a density of 1.26 g/ml, whereas extracellular enveloped arisen by recent gene duplication. More likely, they were virus (EEV) has an extra lipid envelope and a buoyant generated by ancient gene duplication, or the genes density of 1.24 g/ml in CsCl. In VV, EEV is very important were acquired independently from the host. The func- for virus dissemination and mutants unable to make EEV tions of these proteins are unknown. are attenuated in vivo. Several proteins specific for the There are several other multigene families in YLDV, for EEV outer envelope are important for the formation or instance, genes 71L and 145R encode chemokine recep- dissemination of EEV. In YLDV, there are proteins related tor-like proteins, genes 3L and 4L encode proteins re- to the VV EEV proteins F13L, A33R, and A34R (genes 27L, lated to each other and to SPV C5L, genes 2L, 128L, 122R, and 123R, respectively), but counterparts of A36R 136R, and 141R encode members of the immunoglobulin and A56R are absent. Another VV EEV-specific protein superfamily, and genes 19L and 140R encode proteins (B5R) is related to complement control proteins. Although with kelch repeats. 182 LEE, ESSANI, AND SMITH

FIG. 5. Amino acid alignment of YLDV ORF 13L with related protein sequences from cells and other poxviruses generated using the ClustalW program. Arrows mark the amino acids of the catalytic triad of monoglyceride lipase enzymes. Dotted underline marks the conserved motif GXSXG. Solid underline marked a putative zinc finger motif. The black and gray shaded regions represent amino acids that are identical or similar, respectively, in at least two of the three sequences. M.G. lipase, monoglyceride lipase.

Monoglyceride lipase Immunomodulators Protein 13L is related (37% identity and 60% similarity) Poxviruses express a wide range of proteins that to mouse monoglyceride lipase, which catalyzes the last counteract the host response to infection (for reviews step in the hydrolysis of stored triglycerides in the adi- see McFadden et al., 1995; Smith et al., 1997), and many of pocyte (Karlsson et al., 1997). There is a related but these are clustered in the terminal regions of the genome. slightly shorter ORF in CPV strain GRI-90 (Shchelkunov et YLDV also encodes several proteins that are likely to mod- al., 1998) (Fig. 5), and in VV the gene is disrupted into two ulate the host response to infection (Table 3). smaller ORFs, K5L and K6L. The amino acid alignment Near the left end of the genome, gene 2L encodes a (Fig. 5) shows that the three residues of the proposed protein with amino acid similarity to major histocompat- catalytic triad [S122, in a GXSXG motif (underlined with ibility class I (MHC-I) antigens from several species, but dots), D239, and H269 (arrows)] were all conserved in is most closely related to chicken MHC-I and to a related YLDV 13L, but in CPV D239 is substituted by N. There is protein from swinepox virus (SPV) (Massung et al., 1993). also a zinc finger domain near the N-terminus that is Although the amino acid identity (22%) and similarity conserved in all these proteins (underlined). The enzy- (44%) with chicken MHC-I are low, the YLDV protein matic activity of this protein and its role in the poxvirus contains a predicted signal sequence and residues char- biology would be interesting to study. acteristic of the immunoglobulin (Ig) domain (Fig. 6) and

TABLE 3 YLDV Potential Immunomodulators

YLDV protein Putative mechanism Virus or cellular related proteins

2L Blockage of CTL-induced apoptosis Chicken MHC-I 7L Binding of chemokines Human 7TM-CCR8, CAPV Q2/3L, SPV K2R 10L Inhibition of apoptosis MYX SERP-I 12L Inhibition of eIF2␣ phosphorylation VV K3L, VAR C3L, SFV 008.2L/R 14L IL-18-binding protein SPV C9L, MCV MC053L and MC054L, human and mouse IL-18-binding proteins 16L Inhibition of apoptosis MYX M11L 34L Blockage of dsRNA-induced PKR and 2Ј,5Ј- VV E3L, VAR E3L, MYX M029L, SFV S029L oligoadenylate synthetase activation 128L Affects calcium influx into cells Human CD47, VV A38L, SFV S128L, MYX M128L 133L Synthesizing steroids (3␤-HSD) VV A44L 134R Similar to mda-7/IL-10 protein family 136R IFN-␣/␤-binding protein VV-WR B18R, VAR B17R 141R Negative signaling of IL-2 and IFN-␥ production Human OX-2, MYX MA58, SFV S141R 144R Complement inhibition Human CD46, VV C21L and B5R 145R Binding of chemokines Human 7TM-CCR8, CAPV Q2/3L, SPV K2R 149R Inhibition of apoptosis CPV crmA, VV B13R, MYX M151R, SFV S151R GENOME SEQUENCE OF YLDV 183

FIG. 6. Amino acid alignment of YLDV protein 2L with SPV C1L and related MHC-I sequences from chicken (C) and mouse (M) generated using the ClustalW program. Solid underline marks the Ig domain, and the boxed residues represent the transmembrane anchor sequence of cellular MHC-I proteins. The black and gray shaded regions represent amino acids that are identical or similar, respectively, in at least three of the four sequences. has the domain structure typical of MHC-I. In both YLDV high. This suggests that these proteins may bind che- and SPV the Ig domain is larger than the cellular coun- mokines and modulate the immune response to infec- terparts (Fig. 6). Unlike the membrane-bound MHC-I an- tion. tigens of cells and other viruses, such as human cyto- YLDV encodes two proteins with similarity to serine megalovirus (Browne et al., 1990) and MCV (Senkevich protease inhibitors (serpins) and related proteins have and Moss, 1998), the YLDV protein lacks a C-terminal been described in the other poxviruses (for review see hydrophobic membrane anchor and so might be se- Turner et al., 1995). Protein 10L is most closely related to creted. The 2L ORF spans the junction of the left ITR and the secreted serpin encoded by MYX (MYX SERP-I) and terminates 21 nucleotides into the ITR. ITRs may in- which has been shown to inhibit the activity of several crease or decrease in length as a result of transpositions serine proteases (Nash et al., 1998). Protein 149R has of DNA from one end of the genome to another (Moyer et 38% amino acid identity with CPV crmA (VV B13R). This al., 1980) and so such a transposition may have trun- protein is an intracellular inhibitor of caspase 1 (Ray et cated the 2L ORF and removed the C-terminal membrane al., 1992; Kettle et al., 1997), which has roles in apoptosis anchor and cytoplasmic tail. Whatever its derivation, the and in the processing of pro-interleukin (IL)-1 and pro- present protein is likely to be secreted and might bind ␤2 IL-18 into IL-1 and IL-18, respectively. A similar role is microglobulin and short peptides. Possibly this complex predicted for the YLDV protein. might bind to T cells and thereby prevent lysis of virus- Gene 15L encodes a putative growth factor with 40% infected cells. amino acid identity with MYX growth factor. The YLDV The proteins encoded by genes 7L and 145R are protein has a signal peptide and the epidermal growth related to each other (44.6% identity and 59.4% similarity) factor signature (CLCKRQFNGKRC) and is likely to func- and to the family of chemokine receptors. These hydro- tion in a manner similar to that of other poxvirus growth phobic proteins have a hydrophilic N-terminus, 7 trans- factors. membrane domains, and a short cytoplasmic domain. YLDV encodes several proteins predicted to interfere ORF 7L has 53% identity and 73% similarity with human with IFN. Proteins 12L and 34L are related to VV proteins CC chemokine receptor 8 (Tiffany et al., 1997) and slightly K3L and E3L, which prevent the antiviral activity of IFN- higher similarity with the corresponding protein from induced proteins PKR and 2Ј,5Ј-ologoadenylate syn- Macaca mulatta (Margulies et al., 1999). ORF 145R has thetase within infected cells (for review see Smith et al., 44% identity and 68% similarity with the same human 1998). In addition, YLDV protein 136R shares 28% amino protein. Genes encoding chemokine receptor-like pro- acid identity with VV B18R (strain WR) and B19R (strain teins have been described in SPV and capripox virus CPN) proteins that function as a soluble and cell surface (Massung et al., 1993; Cao et al., 1995) but the proteins type I IFN-binding protein (Colamonici et al., 1995; Sy- have not been characterized. Compared to other poxvi- mons et al., 1995; Alcamı´ et al., 2000). Like the VV protein, rus immunomodulators, the degree of relatedness of the YLDV protein has a signal peptide followed by three YLDV ORFs 7L and 145R with cellular counterparts is Ig domains. YLDV lacks a protein with similarity to the 184 LEE, ESSANI, AND SMITH

FIG. 7. Amino acid alignment of YLDV protein 14L with mouse and human IL-18-binding proteins and related proteins from ectromelia virus (EV) strains Hampstead and Naval, MCV MC053L and MCV054L, and SPV C9L. The alignment was generated using the ClustalW program. The black and gray shaded regions represent amino acids that are identical or similar, respectively, in at least six of the eight sequences. Underlined regions represent hydrophobic signal peptides and boxes indicate potential attachment sites for N-linked glycans. soluble IFN-␥-binding protein made by VV and many A44L, a 3␤-hydroxysteroid dehydrogenase (3␤-HSD) en- other poxviruses. Finally, YLDV protein 14L has similarity zyme. In cells this enzyme is essential for the synthesis to IL-18-binding proteins from humans, mice, and poxvi- of all classes of steroid hormones, mineralocorticoids, ruses (Novick et al., 1999; Xiang and Moss, 1999a, b; glucocorticoids, and sex hormones. The VV protein is Born et al., 2000; Smith et al., 2000) (Fig. 7). Each protein active and contributes to virus virulence (Moore and has a signal peptide (underlined) and several potential Smith, 1992). Potentially, the YLDV protein may function sites for attachment of N-linked glycans (boxed). IL-18, in a similar way. also known as IFN-␥-inducing factor, is a potent cytokine YLDV protein 134R has similarity to melanoma differ- whose activities include the induction of IFN and the entiation antigen (mda)-7 (Jiang et al., 1996) and mem- Th1-type immune response. bers of the IL-10 family (Vieira et al., 1991). The protein YLDV protein 144R has 30% amino acid identity with contains a signal peptide and a hydrophilic extracellular human CD46 (membrane cofactor protein). CD46 is a domain, but lacks an anchor sequence and so is likely to GPI-anchored protein that regulates complement activity be secreted. Some other viruses such as Epstein–Barr on the cell surface, has four short consensus repeat virus and OV encode IL-10-like proteins; however, these (SCR) domains typical of complement regulatory pro- proteins are very closely related to the IL-10 proteins of teins, and is also a cellular receptor for measles virus. their host (human and sheep, respectively), with over The YLDV protein 144R has a signal peptide followed by 80% amino acid identity in each case (Hsu et al., 1990; an extracellular domain containing three SCR domains. Fleming et al., 1997). In contrast, the YLDV protein has At the C-terminus there is a hydrophobic transmembrane only 23% identity and 53% similarity with human IL-10. domain, such that the protein might be retained on the Recently, an IL-10-like protein was discovered in herpes cell surface as a glycoprotein with type I membrane virus simairi (K55), which shares 25% amino acid identity topology. Gene 144R encodes the only YLDV protein with IL-10 (Knappe et al., 2000), and other members of related to complement control proteins, whereas VV has this family are being identified. Therefore, YLDV 134R two proteins: C21L and B5R. C21L is a secreted protein may represent a virus version of another member of this called vaccinia complement protein, which binds C3b family. Melanoma differentiation antigen-7 has anti-pro- and C4b and prevents activation of the complement liferative properties for cancer cells (Jiang et al., 1995, cascade (Kotwal and Moss, 1988; Kotwal et al., 1990). 1996). The potential role of mda-7 in YLDV biology is not However, B5R forms part of the outer envelope of EEV clear. and has not been shown to possess anti-complement YLDV protein 141R has 33% amino acid identity with activity (Engelstad et al., 1992; Isaacs et al., 1992). Al- the human T cell costimulatory protein OX-2 (Mc- though VV does not express a protein with CD46-like Caughan et al., 1987) and to related proteins in SFV and properties, during VV morphogenesis CD46 and other MYX (Cameron et al., 1999; Willer et al., 1999). Some host complement regulatory proteins such as CD55 and herpes viruses also contain OX-2-related proteins (Russo CD59 are incorporated onto the EEV outer envelope et al., 1996). OX-2 is a membrane-bound protein on the (Vanderplasschen et al., 1998). CD55 in particular con- surface of dendritic cells and has two Ig domains tributes to the resistance of EEV to destruction by com- (Gorczynski et al., 1999; Ragheb et al., 1999). In contrast, plement. the sequence of YLDV 141R predicts a soluble protein YLDV protein 133L has 44% amino acid identity with VV with only a single Ig domain (Fig. 8). Potentially, this GENOME SEQUENCE OF YLDV 185

the central region of the genome with a role in transcription (VV D12L), an IMV protein (D13L), an EEV-specific protein (VV F13L), and an immunomodulator (VV E3L) (Fig. 9). In cases where a SPV protein is available for comparison, the YLDV protein is most closely related to SPV. In addition, in the left end of the genome, which has been sequenced in SPV (Massung et al., 1993), the YLDV proteins show a consistently higher amino acid identity and similarity to SPV proteins than to proteins from other poxviruses (Table 1). Excluding other yatapoxviruses, YLDV proteins are most closely related to leporipoxvirus proteins where SPV sequence data were not available. FIG. 8. Schematic representation of the YLDV protein 141R and human OX-2, rhesus radinovirus (RRV) R15 and MYX MA56. Positions of Conclusion cysteines (S), forming intramolecular disulfide bonds, and predicted sites for attachment of N-glycans are marked. NH2, amino terminus; The genome sequence of YLDV, a yatapoxvirus, has COOH, carboxyl terminus. Circles represent Ig domains of the variable been determined. The genome is the smallest poxvirus (V) or constant 2 (C2) types. that has been sequenced hitherto and contains 151 ORFs. Comparisons with other poxviruses show that YLDV has very similar transcriptional and DNA replica- YLDV protein might affect the function of T-cells in tive enzymes and many structural proteins for both IMV mounting an antiviral response. and EEV particles. YLDV contains some potential immu- Immunomodulators from other poxviruses that are ab- nomodulators found in other poxviruses and others not sent from YLDV include soluble proteins that bind tumor reported elsewhere. Phylogenic analysis shows that ␤ necrosis factor, IL-1 , or CC chemokines. YLDV is more closely related to SPV and leporipoxvi- Finally, TPV was reported to encode a secreted, 38- ruses than to orthopox, molluscipox, and avipox viruses. kDa protein that bound IL-2, IL-5, and IFN-␥ (Essani et al., 1994). The sequence of the YLDV genome provides no MATERIALS AND METHODS obvious candidate for such a protein. While it is possible that YLDV and TPV differ in the expression of this protein, Cells and viruses the sequence of 12.5 and 10.7 kb from the left and right ends, respectively, of the TPV genome showed that YLDV OMK cells were grown in minimum essential medium and TPV are very closely related (Ͼ98.6% nucleotide (MEM) containing 5% fetal bovine serum (FBS). A virus identity) and the predicted proteins were extremely sim- sample labeled simian poxvirus (tanapox virus Davies ilar (data not shown). strain) was obtained from the American Type Culture Collection but after growth, purification, and restriction YLDV-specific proteins analysis of the virus genome, this virus was found to be distinct from TPV and indistinguishable from YLDV Seven YLDV genes (22L, 23L, 28R, 126R, 130L, 131R, (Knight et al., 1989). TPV was then obtained from Joe and 132R) encoded proteins with no significant similarity Esposito (Center for Disease Control, Atlanta, GA). to other proteins in databases. These are located toward Stocks of each virus were grown in OMK cells for 5 days. the genomic termini and show clustering, suggesting Virus present in the infected cells was released by three that some of these might have been acquired simulta- cycles of freeze/thawing and sonication and the infectiv- neously. Protein 28R is predicted to encode a protein ity was determined by plaque assay on OMK cells for 11 with one transmembrane domain. Protein 126R has a days under a semisolid overlay [MEM containing 1.5% putative signal peptide, and protein 130L encodes a (w/v) carboxymethyl cellulose, 2.5% FBS]. After removal putative 21.2-kDa secreted protein. Both 126R and 130L of the overlay, cell monolayers were stained with 0.1% are likely to be glycosylated. (w/v) crystal violet in 15% ethanol. YLDV was purified from cytoplasmic extracts of in- Phylogeny fected cells by sucrose density gradient centrifugation as Comparison of YLDV proteins with proteins from other described for VV (Mackett et al., 1985). Briefly, infected poxviruses indicates that YLDV is very closely related to cells were swollen in hypotonic buffer and dounce-ho- YMTV (where sequence availability permits comparison). mogenized, and the nuclei were removed by centrifuga- Outside the yatapoxvirus genus, YLDV is most closely tion. The postnuclear supernatant was sonicated and related to SPV and leporipoxviruses than to orthopoxvi- virus was sedimented through a 36% (w/v) sucrose cush- ruses, FPV and MCV. Unrooted phylogenic trees were ion. This virus pellet was resuspended by sonication and constructed for several proteins, including a protein from trypsinized, and virions were purified by two cycles of 186 LEE, ESSANI, AND SMITH

FIG. 9. Unrooted phylogenic trees of four YLDV proteins and related proteins from other poxviruses. (A) mRNA capping enzyme small subunit (VV D12L). (B) IMV protein (VV D13L). (C) EEV-specific protein (VV F13L). (D) dsRNA-binding protein (VV E3L). The consensus tree was generated as follows: Protein sequences were retrieved from the SWALL protein database and aligned by CLUSTAL W. Data were processed by PRODIST and NEIGHBOR (which generated 100 different phylogenic trees), and finally CONSENSUS was used to draw the consensus trees. OV, orf virus. centrifugation in 15–40% (w/v) sucrose density gradients. of approximately 10 kb were ligated into BamHI-digested Virus was stored in 1 mM Tris–HCl (pH 9). Virus DNA pUC118. Each library of ligated fragments was trans- was extracted from purified virus as described (Esposito formed into Escherichia coli TG1 cells and bacterial et al., 1981). clones were selected on agar plates containing ampicil- lin. In some cases, bacteriophage m13K07 was used to Construction of sequencing library and DNA infect pUC118-transformed TG1 cells to make single- sequencing stranded (ss)DNA as described by the manufacturer Virus DNA was broken randomly by sonication, and (New England Biolabs). DNA (ss or ds) for sequencing DNA fragments were made blunt-ended by treatment reactions was prepared by a Qiagen BioRobot 9600. with S1 nuclease (Gibco BRL) and resolved by agarose Clones were sequenced using the dideoxy chain ter- gel electrophoresis. Fragments of 0.8 to 2 kb were re- mination method of Sanger et al. (1977) on an ABI 373 covered by electroelution and ligated into SmaI-digested automated DNA sequencer using cycle sequencing and pUC118 DNA that had been treated with calf intestinal dye terminators (ABI Prism BigDye Termination Cycle alkaline phosphatase to remove 5Ј-phosphate groups. To Sequencing Ready Reaction Kit with AmpliTaq DNA Poly- obtain larger DNA fragments for primer walking, virus merase). Random DNA sequence was processed with DNA was digested partially with Sau3AI and fragments the Staden computer program package (Bonfield et al., GENOME SEQUENCE OF YLDV 187

1995). The program PREGAP was used to convert data Altschul, S. F., Gish, W., Miller, W., Myers, E. W., and Lipman, D. J. (1990). into a format suitable for assembly, to identify vector Basic local alignment search tool. J. Mol. Biol. 215, 403–410. sequence, and to eliminate poor-quality data. Base call- Amano, A., and Miyamura, T. (2000). DNA sequence of Yaba virus BamHI-D fragment. DDBJ/EMBL/GenBank databases. ing and quality checks were performed manually using Amano, H., Kato, K., and Miyamura, T. (2000a). Sequence analysis of TREV and contaminating host sequences were identified Yaba virus DNA. DDBJ/EMBL/GenBank databases. and eliminated using REPE. Assembly was achieved Amano, H., Morikawa, S., Ueda, Y., and Miyamura, T. (2000b). Nucleo- with GAP4. During the final stages of assembly, gaps tide sequence of the central 50-kbp region of Yaba virus DNA. were filled by sequencing directly from whole virus DNA DDBJ/EMBL/GenBank databases. Amano, H., Ueda, Y., and Miyamura, T. (1995). Identification and char- using oligonucleotide primers. acterization of the thymidine kinase gene of Yaba virus. J. Gen. Virol. Determination of the DNA sequence composition and 76, 1109–1115. identification of ORFs were achieved using the program Amegadzie, B. Y., Ahn, B. Y., and Moss, B. (1991). Identification, se- XNIP (Bonfield et al., 1995). Sequence similarity searches quence, and expression of the gene encoding a Mr 35,000 subunit of with the protein database SWALL (European Bioinformat- the vaccinia virus DNA-dependent RNA polymerase. J. Biol. Chem. 266, 13712–13718. ics Institute, EBI, http://www.ebi.ac.uk) were performed Amegadzie, B. Y., Ahn, B. Y., and Moss, B. (1992). Characterization of a with WU-BLASP (Altschul et al., 1990; Altschul and Gish, 7-kilodalton subunit of vaccinia virus DNA-dependent RNA polymer- 1996). Programs XEGCG (Program manual for the Wis- ase with structural similarities to the smallest subunit of eukaryotic consin Package, Version 8, Genetics Computer Group, RNA polymerase II. J. Virol. 66, 3003–3010. Madison, WI) and SMART version 2.0 (Schultz et al., Antoine, G., Scheiflinger, F., Dorner, F., and Falkner, F. G. (1998). The complete genomic sequence of the modified vaccinia Ankara strain: 1998) were used for detailed analysis of each protein. Comparison with other orthopoxviruses. Virology 244, 365–396. Programs ClustalW (Thompson et al., 1994) and Genedoc Baldick, C. J., Keck, J. G., and Moss, B. (1992). Mutational analysis of the (version 2.0) (http://www.cris.com/ϳketchup/genedoc. core, spacer, and initiator regions of vaccinia virus intermediate- shtml) were used to manipulate the multiple sequence class promoters. J. Virol. 66, 4710–4719. file. TreeView (Page, 1996) was used for phylogenetic Baroudy, B. M., and Moss, B. (1982). Sequence homologies of diverse length tandem repetitions near ends of vaccinia virus genome sug- analysis. gest unequal crossing over. Nucleic Acids Res. 10, 5673–5679. Baroudy, B. M., Venkatesan, S., and Moss, B. (1982). Incompletely ACKNOWLEDGMENTS base-paired flip-flop terminal loops link the two DNA strands of the vaccinia virus genome into one uninterrupted polynucleotide chain. This work was supported by a program grant from the United King- Cell 28, 315–324. dom Medical Research Council and support from the EPA Cephalo- Bayliss, C. D., and Smith, G. L. (1996). Vaccinia virion protein I8R has sporin Trust. We thank Joe Esposito (CDC, Atlanta, GA) for advice and both DNA and RNA helicase activities: Implications for vaccinia virus help in providing yatapoxviruses, the Oxford University Bioinformatics transcription. J. Virol. 70, 794–800. staff for computational assistance, and Jeremy Gardner for critical Bayliss, C. D., and Smith, G. L. (1997). Vaccinia virion protein VP8, the reading of the manuscript. 25 kDa product of the L4R gene, binds single-stranded DNA and RNA with similar affinity. Nucleic Acids Res. 25, 3984–3990. REFERENCES Bearcroft, W. G., and Jamieson, M. F. (1958). An outbreak of subcutaneous tumors in rhesus monkeys. Nature 182, 195–196. Afonso, C. L., Tulman, E. R., Lu, Z., Oma, E., Kutish, G. F., and Rock, D. L. Beattie, E., Tartaglia, J., and Paoletti, E. (1991). Vaccinia-virus encoded (1999). The genome of Melanoplus sanguinipes entomopoxvirus. eIF-2␣ homolog abrogates the antiviral effect of interferon. Virology J. Virol. 73, 533–552. 183, 419–422. Afonso, C. L., Tulman, E. R., Lu, Z., Zsak, L., Kutish, G. F., and Rock, D. L. Bertholet, C., Stocco, P., Van Meir, E., and Wittek, R. (1986). Functional (2000). The genome of fowlpox virus. J. Virol. 74, 3815–3831. analysis of the 5Ј flanking sequence of a vaccinia virus late gene. Ahn, B. Y., Gershon, P. D., Jones, E. V., and Moss, B. (1990a). Identifi- EMBO J. 5, 1951–1957. cation of rpo30, a vaccinia virus RNA polymerase gene with struc- Betakova, T., Wolffe, E. J., and Moss, B. (2000). The vaccinia virus A14.5L tural similarity to a eucaryotic transcription elongation factor. Mol. gene encodes a hydrophobic 53-amino-acid virion membrane pro- Cell. Biol. 10, 5433–5441. tein that enhances virulence in mice and is conserved among ver- Ahn, B. Y., Jones, E. V., and Moss, B. (1990b). Identification of the tebrate poxviruses. J. Virol. 74, 4085–4092. vaccinia virus gene encoding an 18-kilodalton subunit of RNA poly- Binns, M. M., Stenzler, L., Tomley, F. M., Campbell, J., and Boursnell, merase and demonstration of a 5Ј poly(A) leader on its early tran- M. E. (1987). Identification by a random sequencing strategy of the script. J. Virol. 64, 3019–3024. fowlpoxvirus DNA polymerase gene, its nucleotide sequence and Ahn, B. Y., and Moss, B. (1992). RNA polymerase-associated transcrip- comparison with other viral DNA polymerases. Nucleic Acids Res. tion specificity factor encoded by vaccinia virus. Proc. Natl. Acad. Sci. 15, 6563–6573. USA 89, 3536–3540. Bonfield, J. K., Smith, K., and Staden, R. (1995). A new DNA sequence Ahn, B. Y., Rosel, J., Cole, N. B., and Moss, B. (1992). Identification and assembly program. Nucleic Acids Res. 23, 4992–4999. expression of rpo19, a vaccinia virus gene encoding a 19-kilodalton Boone, R. F., and Moss, B. (1978). Sequence complexity and relative DNA-dependent RNA polymerase subunit. J. Virol. 66, 971–982. abundance of vaccinia virus mRNAs synthesized in vivo and in vitro. Alcamı´, A., Symons, J. A., and Smith, G. L. (2000). The vaccinia virus J. Virol. 26, 554–569. soluble alpha/beta interferon (IFN) receptor binds to the cell surface Born, T. L., Morrison, L. A., Esteban, D. J., VandenBos, T., Thebeau, L. G., and protects cells from the antiviral effects of IFN. J. Virol. 74, Chen, N., Spriggs, M. K., Sims, J. E., and Buller, R. M. L. (2000). A 11230–11239. poxvirus proteins that binds to and inactivates IL-18, and inhibits NK Altschul, S. F., and Gish, W. (1996). Local alignment statistics. In “Meth- cell response. J. Immunol. 164, 3246–3254. ods in Enzymology” (R. Doolittle, Ed.), Vol. 266, pp. 460–480. Aca- Boursnell, M. E. G., Foulds, I. J., Campbell, J. I., and Binns, M. M. (1988). demic Press, San Diego. Non-essential genes in the vaccinia virus HindIII K fragment: A gene 188 LEE, ESSANI, AND SMITH

related to serine protease inhibitors and a gene related to the 37K G., and Krijnse Locker, J. (1996). A vaccinia virus core protein, p39, is vaccinia virus major envelope antigen. J. Gen. Virol. 69, 2995–3003. membrane associated. J. Virol. 70, 6909–6921. Brick, D. J., Burke, R. D., Schiff, L., and Upton, C. (1998). Shope fibroma Davis, R. E., and Mathews, C. K. (1993). Acidic C terminus of vaccinia virus RING finger protein N1R binds DNA and inhibits apoptosis. virus DNA-binding protein interacts with ribonucleotide reductase. Virology 249, 42–51. Proc. Natl. Acad. Sci. USA 90, 745–749. Browne, H., Smith, G. L., Beck, S., and Minson, T. (1990). A complex Davison, A. J., and Moss, B. (1989). Structure of vaccinia virus late between the MHC class I homologue encoded by human cytomeg- promoters. J. Mol. Biol. 210, 771–784. alovirus and beta 2 microglobulin. Nature 347, 770–772. deHarven, E., and Yohn, D. S. (1966). The fine structure of the Yaba Broyles, S. S. (1993). Vaccinia virus encodes a functional dUTPase. monkey tumor poxvirus. Cancer Res. 26, 995–1008. Virology 195, 863–865. Deng, L., and Shuman, S. (1998). Vaccinia NPH-I, a DExH-box ATPase, Broyles, S. S., and Fesler, B. S. (1990). Vaccinia virus gene encoding a is the energy coupling factor for mRNA transcription termination. component of the viral early transcription factor. J. Virol. 64, 1523– Genes Dev. 12, 538–546. 1529. Douglas, N. J., and Dumbell, K. R. (1996). DNA sequence variation as a Broyles, S. S., and Moss, B. (1986). Homology between RNA poly- clue to the phylogenesis of orthopoxviruses. J. Gen. Virol. 77, 947– merases of poxviruses, prokaryotes, and eukaryotes: Nucleotide 951. sequence and transcriptional analysis of vaccinia virus genes en- Downie, A. W., and Espanã, C. (1972). Comparison of Tanapox virus and coding 147-kDa and 22-kDa subunits. Proc. Natl. Acad. Sci. USA 83, Yaba-like viruses causing epidemic disease in monkeys. J. Hyg. 70, 3141–3145. 23–32. Cameron, C., Hota-Mitchell, S., Chen, L., Barrett, J., Cao, J. X., Macaulay, Downie, A. W., Taylor Robinson, C. H., Caunt, A. E., Nelson, G. S., C., Willer, D., Evans, D., and McFadden, G. (1999). The complete DNA Manson Bahr, P. E., and Matthews, T. C. (1971). Tanapox: A new sequence of myxoma virus. Virology 264, 298–318. disease caused by a pox virus. Br. Med. J. 1, 363–368. Campbell, I. G., Freemont, P. S., Foulkes, W., and Trowsdale, J. (1992). Duncan, S. A., and Smith, G. L. (1992). Identification and characteriza- An ovarian tumor marker with homology to vaccinia virus contains an tion of an extracellular envelope glycoprotein affecting vaccinia virus IgV-like region and multiple transmembrane domains. Cancer Res. egress. J. Virol. 66, 1610–1621. 52, 5416–5120. Dyster, L. M., and Niles, E. G. (1991). Genetic and biochemical characterization of vaccinia virus genes D2L and D3R which encode virion Cao, J. X., Gershon, P. D., and Black, D. N. (1995). Sequence analysis of structural proteins. Virology 182, 455–467. HindIII Q2 fragment of capripoxvirus reveals a putative gene encod- Earl, P. L., Jones, E. V., and Moss, B. (1986). Homology between DNA ing a G-protein-coupled chemokine receptor homologue. Virology polymerases of poxviruses, herpesviruses, and adenoviruses: Nu- 209, 207–212. cleotide sequence of the vaccinia virus DNA polymerase gene. Proc. Casey, H. W., Woodruff, J. M., and Butcher, W. I. (1967). Electron micros- Natl. Acad. Sci. USA 83, 3659–3663. copy of a benign epidermal pox disease of rhesus monkeys. Am. J. Engelstad, M., Howard, S. T., and Smith, G. L. (1992). A constitutively Pathol. 51, 431–446. expressed vaccinia gene encodes a 42-kDa glycoprotein related to Cassetti, M. C., Merchlinsky, M., Wolffe, E. J., Weisberg, A. S., and Moss, complement control factors that forms part of the extracellular virus B. (1998). DNA packaging mutant: Repression of the vaccinia virus envelope. Virology 188, 801–810. A32 gene results in noninfectious, DNA-deficient, spherical, envel- Espanã, C. (1971). A pox disease of monkeys transmittal to man. In oped particles. J. Virol. 72, 5769–5780. “Medical Primatology” (E. I. Goldsmith and J. Moor-Jankowski, Eds.), Chang, H. W., Uribe, L. H., and Jacobs, B. L. (1995). Rescue of vaccinia pp. 694–708. S. Karger, Basel. Proceedings of the 2nd conference on virus lacking the E3L gene by mutants of E3L. J. Virol. 69, 6605–6608. Experimental Medicine and Surgery in Primates. Chang, H. W., Watson, J. C., and Jacobs, B. L. (1992). The E3L gene of Espanã, C., Brayton, M. A., and Ruebner, B. H. (1971). Electron micros- vaccinia virus encodes an inhibitor of the interferon-induced, double- copy of the Tana poxvirus. Exp. Mol. Pathol. 15, 34–42. stranded RNA-dependent protein kinase. Proc. Natl. Acad. Sci. USA Esposito, J. R., Condit, R. C., and Obijeski, J. (1981). The preparation of 89, 4825–4829. orthopoxvirus DNA. J. Virol. Meth. 2, 175–179. Chertov, O., Telezhinskaya, I. N., Zaitseva, E. V., Golubeva, T. B., Essani, K., Chalasani, S., Eversole, R., Beuving, L., and Birmingham, L. Zinov’ev, V. V., Ovechkina, L. G., Mazkova, L. B., and Malygin, E. G. (1994). Multiple anti-cytokine activities secreted from tanapox virus- (1991). Amino acid sequence determination of vaccinia virus immu- infected cells. Microb. Pathog. 17, 347–353. nodominant protein p35 and identification of the gene. Biomed. Sci. Evans, E., Klemperer, N., Ghosh, R., and Traktman, P. (1995). The 2, 151–154. vaccinia virus D5 protein, which is required for DNA replication, is a Child, S. J., Palumbo, G. J., Buller, R. M. L., and Hruby, D. E. (1990). nucleic acid-independent nucleoside triphosphatase. J. Virol. 69, Insertional inactivation of the large subunit of ribonucleotide reduc- 5353–5361. tase encoded by vaccinia virus is associated with reduced virulence Fenner, F. (1996). Poxviruses. In “Fields Virology” (B. N. Fields, D. M. in vivo. Virology 174, 625–629. Knipe, and P. M. Howley, Eds.), 3rd ed., Vol. 2, pp. 2673–2702. Christen, L. M., Sanders, M., Wiler, C., and Niles, E. G. (1998). Vaccinia Lippincott–Raven, Philadelphia. virus nucleoside triphosphate phosphohydrolase I is an essential Fleming, S. B., McCaughan, C. A., Andrews, A. E., Nash, A. D., and viral early gene transcription termination factor. Virology 245, 360– Mercer, A. A. (1997). A homolog of interleukin-10 is encoded by the 371. poxvirus orf virus. J. Virol. 71, 4857–4861. Chung, C. S., Hsiao, J. C., Chang, Y. S., and Chang, W. (1998). A27L Forget, B. G., Tse, W. T., Gallagher, P. G., Scarpa, A. L., and Lux, S. E. protein mediates vaccinia virus interaction with cell surface heparan (1997). Human ankyrin (ANK1) gene, exon 42 and complete cds. sulfate. J. Virol. 72, 1577–1585. DDBJ/EMBL/Genbank databases (GeneBank accession No. Colamonici, O. R., Domanski, P., Sweitzer, S. M., Larner, A., and Buller, Q99407). R. M. L. (1995). Vaccinia virus B18R gene encodes a type I interferon- Franke, C. A., Wilson, E. M., and Hruby, D. E. (1990). Use of a cell-free binding protein that blocks interferon ␣ transmembrane signaling. system to identify the vaccinia virus L1R gene product as the major J. Biol. Chem. 270, 15974–15978. late myristylated virion protein M25. J. Virol. 64, 5988–5996. Condit, R. C., Xiang, Y., and Lewis, J. I. (1996). Mutation of vaccinia virus Gershon, P. D., Ahn, B. Y., Garfield, M., and Moss, B. (1991). Poly(A) gene G2R causes suppression of gene A18R ts mutants: Implications polymerase and a dissociable polyadenylation stimulatory factor for control of transcription. Virology 220, 10–19. encoded by vaccinia virus. Cell 66, 1269–1278. Cudmore, S., Blasco, R., Vincentelli, R., Esteban, M., Sodeik, B., Griffiths, Gershon, P. D., Ansell, D. M., and Black, D. N. (1989). A comparison of GENOME SEQUENCE OF YLDV 189

the genome organization of capripoxvirus with that of the orthopox- brane glycoprotein component of the extracellular virus envelope. viruses. J. Virol. 63, 4703–4708. J. Virol. 66, 7217–7224. Gershon, P. D., and Black, D. N. (1989). The nucleotide sequence Jackson, R. J., and Bults, H. G. (1990). A myxoma virus nucleotide around the capripoxvirus thymidine kinase gene reveals a gene sequence with homology to the vaccinia virus RNA polymerase shared specifically with leporipoxvirus. J. Gen. Virol. 70, 525–533. 22-kDa subunit gene. Nucleic Acids Res. 18, 5290. Gershon, P. D., and Moss, B. (1990). Early transcription factor subunits Jackson, R. J., and Bults, H. G. (1992). The myxoma virus thymidine are encoded by vaccinia virus late genes. Proc. Natl. Acad. Sci. USA kinase gene: Sequence and transcriptional mapping. J. Gen. Virol. 87, 4401–4405. 73, 323–328. Gershon, P. D., and Moss, B. (1993). Stimulation of poly(A) tail elonga- Jackson, R. J., and Hall, D. F. (1998). The myxoma virus EcoRI-O frag- tion by the VP39 subunit of the vaccinia virus-encoded poly(A) poly- ment encodes the DNA binding core protein and the major envelope merase. J. Biol. Chem. 268, 2203–2210. protein of extracellular poxvirus. Virus Genes 17, 55–62. Goebel, S. J., Johnson, G. P., Perkus, M. E., Davis, S. W., Winslow, J. P., Jackson, R. J., Hall, D. F., and Kerr, P. J. (1996). Construction of recom- and Paoletti, E. (1990). The complete DNA sequence of vaccinia binant myxoma viruses expressing foreign genes from different in- virus. Virology 179, 247–266. tergenic sites without associated attenuation. J. Gen. Virol. 77, 1569– Gorczynski, R. M., Cattral, M. S., Chen, Z., Hu, J., Lei, J., Min, W. P., Yu, 1575. G., and Ni, J. (1999). An immunoadhesin incorporating the molecule Jackson, R. J., Hall, D. F., and Kerr, P. J. (1999). Myxoma virus encodes ␣ OX-2 is a potent immunosuppressant that prolongs allo- and xeno- an -2,3-sialyltransferase that enhances virulence. J. Virol. 73, 2376– graft survival. J. Immunol. 163, 1654–1660. 2384. Graham, K. A., Opgenorth, A., Upton, C., and McFadden, G. (1992). Jensen, O. N., Houthaeve, T., Shevchenko, A., Cudmore, S., Ashford, T., Myxoma virus M11L ORF encodes a protein for which cell surface Mann, M., Griffiths, G., and Krijnse Locker, J. (1996). Identification of localization is critical in manifestation of viral virulence. Virology 191, the major membrane and core proteins of vaccinia virus by two- 112–124. dimensional electrophoresis. J. Virol. 70, 7485–7497. Guan, K. L., Broyles, S. S., and Dixon, J. E. (1991). A Tyr/Ser protein Jiang, H., Lin, J. J., Su, Z. Z., Goldstein, N. I., and Fisher, P. B. (1995). phosphatase encoded by vaccinia virus. Nature 350, 359–362. Subtraction hybridization identifies a novel melanoma differentiation Gubser, C., and Smith, G. L. Camelpox and smallpox: Too close for associated gene, mda-7, modulated during human melanoma differentiation, growth and progression. Oncogene 11, 2477–2486. comfort? Submitted for publication. Jiang, H., Su, Z. Z., Lin, J. J., Goldstein, N. I., Young, C. S., and Fisher, P. B. Gvakharia, B. O., Koonin, E. K., and Mathews, C. K. (1996). Vaccinia virus (1996). The melanoma differentiation associated gene mda-7 sup- G4L gene encodes a second glutaredoxin. Virology 226, 408–411. presses cancer cell growth. Proc. Natl. Acad. Sci. USA 93, 9160–9165. Haig, D. M., McInnes, C. J., Thomson, J., Wood, A., Bunyan, K., and Kane, E. M., and Shuman, S. (1992). Temperature-sensitive mutations in Mercer, A. (1998). The orf virus OV20.0L gene product is involved in the vaccinia virus H4 gene encoding a component of the virion RNA interferon resistance and inhibits an interferon-inducible, double- polymerase. J. Virol. 66, 5752–5762. stranded RNA-dependent kinase. Immunology 93, 335–340. Kane, E. M., and Shuman, S. (1993). Vaccinia virus morphogenesis is Hall, A. S., and McNulty, W. P. (1967). A contagious pox disease in blocked by a temperature-sensitive mutation in the I7 gene that monkeys. J. Am. Vet. Med. Assoc. 151, 833–838. encodes a virion component. J. Virol. 67, 2689–2698. Hertig, C., Coupar, B. E., Gould, A. R., and Boyle, D. B. (1997). Field and Kao, S. Y., and Bauer, W. R. (1987). Biosynthesis and phosphorylation of vaccine strains of fowlpox virus carry integrated sequences from the vaccinia virus structural protein VP11. Virology 159, 399–407. avian retrovirus, reticuloendotheliosis virus. Virology 235, 367–376. Karlsson, M., Contreras, J. A., Hellman, U., Tornqvist, H., and Holm, C. Hirt, P., Hiller, G., and Wittek, R. (1986). Localization and fine structure (1997). cDNA cloning, tissue distribution, and identification of the of a vaccinia virus gene encoding an envelope antigen. J. Virol. 58, catalytic triad of monoglyceride lipase. Evolutionary relationship to 757–764. esterases, lysophospholipases, and haloperoxidases. J. Biol. Chem. Howard, S. T., Chan, Y. S., and Smith, G. L. (1991). Vaccinia virus 272, 27218–27223. homologues of the Shope fibroma virus inverted terminal repeat Keck, J. G., Baldick, C. J., Jr., and Moss, B. (1990). Role of DNA replica- proteins and a discontinuous ORF related to the tumor necrosis tion in vaccinia virus gene expression: A naked template is required factor receptor family. Virology 180, 633–647. for transcription of three late trans-activator genes. Cell 61, 801–809. Howard, S. T., and Smith, G. L. (1989). Two early vaccinia virus genes Keck, J. G., Kovacs, G. R., and Moss, B. (1993). Overexpression, purifi- encode polypeptides related to protein kinases. J. Gen. Virol. 70, cation, and late transcription factor activity of the 17-kilodalton pro- 3187–3201. tein encoded by the vaccinia virus A1L gene. J. Virol. 67, 5740–5748. Hruby, D. E., Maki, R. A., Miller, D. B., and Ball, L. A. (1983). Fine Kerr, S. M., Johnston, L. H., Odell, M., Duncan, S. A., Law, K. M., and structure analysis and nucleotide sequence of the vaccinia virus Smith, G. L. (1991). Vaccinia DNA ligase complements Saccharomy- thymidine kinase gene. Proc. Natl. Acad. Sci. USA 80, 3411–3415. ces cerevisiae cdc9, localizes in cytoplasmic factories and affects Hsiao, J. C., Chung, C. S., and Chang, W. (1999). Vaccinia virus envelope virulence and virus sensitivity to DNA damaging agents. EMBO J. 10, D8L protein binds to cell surface chondroitin sulfate and mediates 4343–4350. the adsorption of intracellular mature virions to cells. J. Virol. 73, Kettle, S., Alcamı´, A., Khanna, A., Ehret, R., Jassoy, C., and Smith, G. L. 8750–8761. (1997). Vaccinia virus serpin B13R (SPI-2) inhibits interleukin-1␤- Hsu, D. H., de Waal Malefyt, R., Fiorentino, D. F., Dang, M. N., Vieira, P., converting enzyme and protects virus-infected cells from TNF- and de Vries, J., Spits, H., Mosmann, T. R., and Moore, K. W. (1990). Fas-mediated apoptosis, but does not prevent IL-1␤-induced fever. Expression of interleukin-10 activity by Epstein–Barr virus protein J. Gen. Virol. 78, 677–685. BCRF1. Science 250, 830–832. Klemperer, N., Ward, J., Evans, E., and Traktman, P. (1997). The vaccinia Hubbs, A. E., and Wright, C. F. (1996). The A2L intermediate gene virus I1 protein is essential for the assembly of mature virions. J. Virol. product is required for in vitro transcription from a vaccinia virus late 71, 9285–9294. promoter. J. Virol. 70, 327–331. Knappe, A., Hor, S., Wittmann, S., and Fickenscher, H. (2000). Induction Hunt, H. D., and Fulton, J. E. (1998). Analysis of polymorphisms in the of a novel cellular homolog of interleukin-10, AK155, by transforma- major expressed class I locus (B-FIV) of the chicken. Immunogenet- tion of T lymphocytes with herpesvirus saimiri. J. Virol. 74, 3881–3887. ics 47, 456–467. Knight, J. C., Novembre, F. J., Brown, D. R., Goldsmith, C. S., and Isaacs, S. N., Wolffe, E. J., Payne, L. G., and Moss, B. (1992). Charac- Esposito, J. J. (1989). Studies on tanapox virus. Virology 172, 116–124. terization of a vaccinia virus-encoded 42-kilodalton class I mem- Koonin, E. V. (1993). A highly conserved sequence motif defining the 190 LEE, ESSANI, AND SMITH

family of MutT-related proteins from eubacteria, eukaryotes and erratum appears in EMBO J. 1992 Sep; 11(9):3490]. EMBO J. 11, viruses. Nucleic Acids Res. 21, 4847. 1973–1980. Koonin, E. V., Senkevich, T. G., and Chernos, V. I. (1993). Gene A32 Morgan, J. R., Cohen, L. K., and Roberts, B. E. (1984). Identification of the product of vaccinia virus may be an ATPase involved in viral DNA DNA sequences encoding the large subunit of the mRNA-capping packaging as indicated by sequence comparisons with other puta- enzyme of vaccinia virus. J. Virol. 52, 206–214. tive viral ATPases. Virus Genes 7, 89–94. Moss, B. (1996). Poxviridae: The viruses and their replication. In “Fields Kotwal, G. J., Isaacs, S. N., McKenzie, R., Frank, M. M., and Moss, B. Virology” (B. N. Fields, D. M. Knipe, and P. M. Howley, Eds.), 3rd ed., (1990). Inhibition of the complement cascade by the major secretory Vol. 2, pp. 2637–2671. Lippincott–Raven, Philadelphia. protein of vaccinia virus. Science 250, 827–830. Mossman, K., Lee, S. F., Barry, M., Boshkov, L., and McFadden, G. Kotwal, G. J., and Moss, B. (1988). Vaccinia virus encodes a secretory (1996). Disruption of M-T5, a novel myxoma virus gene member of polypeptide structurally related to complement control proteins. Na- poxvirus host range superfamily, results in dramatic attenuation of ture 335, 176–178. myxomatosis in infected European rabbits. J. Virol. 70, 4394–4410. Kovacs, G. R., and Moss, B. (1996). The vaccinia virus H5R gene Moyer, R. W., Graves, R. L., and Rothe, C. T. (1980). The white pock (␮) encodes late gene transcription factor 4: Purification, cloning, and mutants of rabbit poxvirus, III. Terminal DNA sequence duplication overexpression. J. Virol. 70, 6796–6802. and transposition in rabbit poxvirus. Cell 22, 545–553. Kupper, J. L., Casey, H. W., and Johnson, D. K. (1970). Experimental Yaba Nash, P., Whitty, A., Handwerker, J., Macen, J., and McFadden, G. (1998). and benign epidermal monkey pox in rhesus monkeys. Lab. Anim. Inhibitory specificity of the anti-inflammatory myxoma virus serpin, Care 20, 979–988. SERP-1. J. Biol. Chem. 273, 20982–20991. Lin, C. L., Chung, C. S., Heine, H. G., and Chang, W. (2000). Vaccinia Neering, J., and Essani, K. (1999). Tanapox virus 2.1kbp PstI-L fragment. virus envelope H3L protein binds to cell surface heparan sulfate and DDBJ/EMBL/GenBank databases. is important for intracellular mature virion morphogenesis and virus Nezu, J., Oku, A., Jones, M. H., and Shimane, M. (1997). Identification of infection in vitro and in vivo. J. Virol. 74, 3353–3365. two novel human putative serine/threonine kinases, VRK1 and VRK2, Lin, S., and Broyles, S. S. (1994). Vaccinia protein kinase 2: A second with structural similarity to vaccinia virus B1R kinase. Genomics 45, essential serine/threonine protein kinase encoded by vaccinia virus. 327–331. Proc. Natl. Acad. Sci. USA 91, 7653–7657. Nicholas, A. H., and McNulty, W. P. (1968). In vitro characteristics of a poxvirus isolated from rhesus monkeys. Nature 217, 745–746. Liszewski, M. K., Post, T. W., and Atkinson, J. P. (1991). Membrane Niles, E. G., Lee-Chen, G. J., Shuman, S., Moss, B., and Broyles, S. S. cofactor protein (MCP or CD46): Newest member of the regulators of (1989). Vaccinia virus gene D12L encodes the small subunit of the complement activation gene cluster. Annu. Rev. Immunol. 9, 431–455. viral mRNA capping enzyme. Virology 172, 513–522. Mackett, M., Smith, G. L., and Moss, B. (1985). The construction and Niles, E. G., and Seto, J. (1988). Vaccinia virus gene D8 encodes a virion characterization of vaccinia virus recombinants expressing foreign transmembrane protein. J. Virol. 62, 3772–3778. genes. In “DNA Cloning: A Practical Approach” (D. M. Glover, Ed.), Niven, J. S. F., Armstrong, J. A., Andrewes, C. H., Pereira, H. G., and Vol. 2, pp. 191–211. IRL Press, Oxford. Valentine, R. C. (1961). Subcutaneous growths in monkeys produced Margulies, B. J., Hauer, D. A., and Clements, J. E. (1999). Identification by poxvirus. J. Pathol. Bacteriol. 81, 1–14. and characterization of thirteen rhesus macaque chemokine recep- Novick, D., Kim, S. H., Fantuzzi, G., Reznikov, L. L., Dinarello, C. A., and tors and chemokine receptor homologues. DDBJ/EMBL/GenBank Rubinstein, M. (1999). Interleukin-18 binding protein: A novel modu- databases. lator of the Th1 cytokine response. Immunity 10, 127–136. Martin, K. H., Grosenbach, D. W., Franke, C. A., and Hruby, D. E. (1997). Oguiura, N., Spehner, D., and Drillien, R. (1993). Detection of a protein Identification and analysis of three myristylated vaccinia virus late encoded by the vaccinia virus C7L open reading frame and study of proteins. J. Virol. 71, 5218–5226. its effect on virus multiplication in different cell lines. J. Gen. Virol. 74, Massung, R. F., Jayarama, V., and Moyer, R. W. (1993). DNA sequence 1409–1413. analysis of conserved and unique regions of swinepox virus: Iden- Page, R. D. (1996). TreeView: An application to display phylogenetic tification of genetic elements supporting phenotypic observations trees on personal computers. Comput. Appl. Biosci. 12, 357–358. including a novel G protein-coupled receptor homologue. Virology Parkinson, J. E., Sanderson, C. M., and Smith, G. L. (1995). The vaccinia 197, 511–528. virus A38L gene product is a 33-kDa integral membrane glycopro- Massung, R. F., Liu, L. I., Qi, J., Knight, J. C., Yuran, T. E., Kerlavage, A. R., tein. Virology 214, 177–188. Parsons, J. M., Venter, J. C., and Esposito, J. J. (1994). Analysis of the Parkinson, J. E., and Smith, G. L. (1994). Vaccinia virus gene A36R complete genome of smallpox variola major virus strain Bangladesh- encodes a M(r) 43–50 K protein on the surface of extracellular 1975. Virology 201, 215–240. enveloped virus. Virology 204, 376–390. McCaughan, G. W., Clark, M. J., and Barclay, A. N. (1987). Character- Patel, D. D., and Pickup, D. J. (1989). The second-largest subunit of the ization of the human homolog of the rat MRC OX-2 membrane poxvirus RNA polymerase is similar to the corresponding subunits of glycoprotein. Immunogenetics 25, 329–335. procaryotic and eucaryotic RNA polymerases. J. Virol. 63, 1076–1086. McDonald, W. F., Klemperer, N., and Traktman, P. (1997). Characteriza- Perkus, M. E., Goebel, S. J., Davis, S. W., Johnson, G. P., Limbach, K., tion of a processive form of the vaccinia virus DNA polymerase. Norton, E. K., and Paoletti, E. (1990). Vaccinia virus host range genes. Virology 234, 168–175. Virology 179, 276–286. McFadden, G., Graham, K., Ellison, K., Barry, M., Macen, J., Schreiber, Petit, F., Bertagnoli, S., Gelfi, J., Fassy, F., Boucraut-Baralon, C., and M., Mossman, K., Nash, P., Lalani, A., and Everett, H. (1995). Inter- Milon, A. (1996). Characterization of a myxoma virus-encoded serpin- ruption of cytokine networks by poxviruses: Lessons from myxoma like protein with activity against interleukin-1 ␤-converting enzyme. virus. J. Leukocyte Biol. 57, 731–738. J. Virol. 70, 5860–5866. McNulty, W. P., Lobitz, W. C., Hu, F., Maruffo, C. A., and Hall, A. S. (1968). Pickup, D. J., Ink, B. S., Parsons, B. L., Hu, W., and Joklik, W. K. (1984). A pox disease in monkeys transmitted to man. Clinical and histolog- Spontaneous deletions and duplications of sequences in the ge- ical features. Arch. Dermatol. 97, 286–293. nome of cowpox virus. Proc. Natl. Acad. Sci. USA 81, 6817–6821. Merchlinsky, M., and Moss, B. (1989). Nucleotide sequence required for Quick, S. D., and Broyles, S. S. (1990). Vaccinia virus gene D7R encodes resolution of the concatemer junction of vaccinia virus DNA. J. Virol. a 20,000-dalton subunit of the viral DNA-dependent RNA polymer- 63, 4354–4361. ase. Virology 178, 603–605. Moore, J. B., and Smith, G. L. (1992). Steroid hormone synthesis by a Ragheb, R., Abrahams, S., Beecroft, R., Hu, J., Ni, J., Ramakrishna, V., Yu, vaccinia enzyme: A new type of virus virulence factor [Published G., and Gorczynski, R. M. (1999). Preparation and functional proper- GENOME SEQUENCE OF YLDV 191

ties of monoclonal antibodies to human, mouse and rat OX-2. Immu- Sequence and genomic analysis of a Rhesus macaque rhadinovirus nol. Lett. 68, 311–315. with similarity to Kaposi’s sarcoma-associated herpesvirus/human Ray, C. A., Black, R. A., Kronheim, S. R., Greenstreet, T. A., Sleath, P. R., herpesvirus 8. J. Virol. 73, 3040–3053. Salvensen, G. S., and Pickup, D. J. (1992). Viral inhibition of inflam- Senkevich, T. G., Bugert, J. J., Sisler, J. R., Koonin, E. V., Darai, G., and mation: Cowpox virus encodes an inhibitor of the interleukin-1␤ Moss, B. (1996). Genome sequence of a human tumorigenic poxvi- converting enzyme. Cell 69, 597–604. rus: Prediction of specific host response-evasion genes. Science Rochester, S. C., and Traktman, P. (1998). Characterization of the single- 273, 813–816. stranded DNA binding protein encoded by the vaccinia virus I3 gene. Senkevich, T. G., Koonin, E. V., Bugert, J. J., Darai, G., and Moss, B. J. Virol. 72, 2917–2926. (1997). The genome of molluscum contagiosum virus: Analysis and Rodriguez, D., Esteban, M., and Rodriguez, J. R. (1995). Vaccinia virus comparison with other poxviruses. Virology 233, 19–42. A17L gene product is essential for an early step in virion morpho- Senkevich, T. G., and Moss, B. (1998). Domain structure, intracellular genesis. J. Virol. 69, 4640–4608. trafficking, and ␤2-microglobulin binding of a major histocompatibil- Rodriguez, J. F., and Smith, G. L. (1990). IPTG-dependent vaccinia virus: ity complex class I homolog encoded by molluscum contagiosum Identification of a virus protein enabling virion envelopment by Golgi virus. Virology 250, 397–407. membrane and egress. Nucleic Acids Res. 18, 5347–5351. Shchelkunov, S. N., Massung, R. F., and Esposito, J. J. (1995). Compar- Roper, R. L., Payne, L. G., and Moss, B. (1996). Extracellular vaccinia ison of the genome DNA sequences of Bangladesh-1975 and India- virus envelope glycoprotein encoded by the A33R gene. J. Virol. 70, 1967 variola viruses. Virus Res. 36, 107–118. 3753–3762. Shchelkunov, S. N., Safronov, P. F., Totmenin, A. V., Petrov, N. A., Rosales, R., Harris, N., Ahn, B. Y., and Moss, B. (1994). Purification and Ryazankina, O. I., Gutorov, V. V., and Kotwal, G. J. (1998). The genomic identification of a vaccinia virus-encoded intermediate stage pro- sequence analysis of the left and right species-specific terminal moter-specific transcription factor that has homology to eukaryotic region of a cowpox virus strain reveals unique sequences and a transcription factor SII (TFIIS) and an additional role as a viral RNA cluster of intact ORFs for immunomodulatory and host range pro- polymerase subunit. J. Biol. Chem. 269, 14260–14267. teins. Virology 243, 432–460. Rosel, J., and Moss, B. (1985). Transcriptional and translational map- Shchelkunov, S. N., Totmenin, A. V., Loparev, V. N., Safronov, P. F., ping and nucleotide sequence analysis of a vaccinia virus gene Gutorov, V. V., Chizhikov, V. E., Knight, J. C., Parsons, J. M., Massung, encoding the precursor of the major core polypeptide 4b. J. Virol. 56, R. F., and Esposito, J. J. (2000). Alastrim smallpox variola minor virus 830–838. genome DNA sequences. Virology 266, 361–386. Rosel, J. L., Earl, P. L., Weir, J. P., and Moss, B. (1986). Conserved Shors, T., Keck, J. G., and Moss, B. (1999). Down regulation of gene TAAATG sequence at the transcriptional and translational initiation expression by the vaccinia virus D10 protein. J. Virol. 73, 791–796. sites of vaccinia virus late genes deduced by structural and func- Shuman, S. (1992). Vaccinia virus RNA helicase: An essential enzyme tional analysis of the HindIII H genome fragment. J. Virol. 60, 436– related to the DE-H family of RNA-dependent NTPases. Proc. Natl. 449. Acad. Sci. USA 89, 10935–10939. Roseman, N. A., and Slabaugh, M. B. (1990). The vaccinia virus HindIII Shuman, S., and Moss, B. (1987). Identification of a vaccinia virus gene F fragment: Nucleotide sequence of the left 6.2 kb. Virology 178, encoding a type I DNA topoisomerase. Proc. Natl. Acad. Sci. USA 84, 410–418. 7478–7482. Ro¨ttger, S., Frischknecht, F., Reckmann, I., Smith, G. L., and Way, M. Simpson, D. A., and Condit, R. C. (1995). Vaccinia virus gene A18R (1999). Interactions between vaccinia virus IEV membrane proteins encodes an essential DNA helicase. J. Virol. 69, 6131–6139. and their roles in IEV assembly and actin tail formation. J. Virol. 73, Smith, G. L., Chan, Y. S., and Howard, S. T. (1991). Nucleotide sequence 2863–2875. of 42 kbp of vaccinia virus strain WR from near the right inverted Rouhandeh, H., Kilpatrick, D., and Vafai, A. (1982). The molecular biol- terminal repeat. J. Gen. Virol. 72, 1349–1376. ogy of Yaba tumour pox virus: Analysis of lipids, proteins and DNA. Smith, G. L., Howard, S. T., and Chan, Y. S. (1989). Vaccinia virus J. Gen. Virol. 62, 207–218. encodes a family of genes with homology to serine proteinase Russo, J. J., Bohenzky, R. A., Chien, M. C., Chen, J., Yan, M., Maddalena, inhibitors. J. Gen. Virol. 70, 2333–2343. D., Parry, J. P., Peruzzi, D., Edelman, I. S., Chang, Y., and Moore, P. S. Smith, G. L., Symons, J. A., and Alcamı´, A. (1998). Poxviruses: Interfering (1996). Nucleotide sequence of the Kaposi sarcoma-associated her- with interferon. Semin. Virol. 8, 409–418. pesvirus (HHV8). Proc. Natl. Acad. Sci. USA 93, 14862–14867. Smith, G. L., Symons, J. A., Khanna, A., Vanderplasschen, A., and Sanderson, C. M., Frischknecht, F., Way, M., Hollinshead, M., and Smith, Alcamı´, A. (1997). Vaccinia virus immune evasion. Immunol. Rev. 159, G. L. (1998). Roles of vaccinia virus EEV-specific proteins in intracel- 137–154. lular actin tail formation and low pH-induced cell–cell fusion. J. Gen. Smith, V. P., Bryant, N. A., and Alcamı´, A. (2000). Ectromelia, vaccinia Virol. 79, 1415–1425. and cowpox viruses encode secreted interleukin-18-binding pro- Sanger, F., Nicklen, S., and Coulson, A. R. (1977). DNA sequencing with teins. J. Gen. Virol. 81, 1223–1230. chain terminating inhibitors. Proc. Natl. Acad. Sci. USA 74, 5463– Spehner, D., Gillard, S., Drillien, R., and Kirn, A. (1988). A cowpox virus 5467. gene required for multiplication in Chinese hamster ovary cells. Sanz, P., and Moss, B. (1999). Identification of a transcription factor, J. Virol. 62, 1297–1304. encoded by two vaccinia virus early genes, that regulates the inter- Strayer, D. S., Jerng, H. H., and O’Connor, K. (1991). Sequence and mediate stage of viral gene expression. Proc. Natl. Acad. Sci. USA analysis of a portion of the genomes of Shope fibroma virus and 96, 2692–2697. malignant rabbit fibroma virus that is important for viral replication in Schmitt, J. F., and Stunnenberg, H. G. (1988). Sequence and transcrip- lymphocytes. Virology 185, 585–595. tional analysis of the vaccinia virus HindIII I fragment. J. Virol. 62, Stuart, D. T., Upton, C., Higman, M. A., Niles, E. G., and McFadden, G. 1889–1897. (1993). A poxvirus-encoded uracil DNA glycosylase is essential for Schnitzlein, W. M., and Tripathy, D. N. (1991). Identification and nucle- virus viability. J. Virol. 67, 2503–2512. otide sequence of the thymidine kinase gene of swinepox virus. Symons, J. A., Alcamı´, A., and Smith, G. L. (1995). Vaccinia virus encodes Virology 181, 727–732. a soluble type I interferon receptor of novel structure and broad Schultz, J., Milpetz, F., Bork, P., and Ponting, C. P. (1998). SMART, a species specificity. Cell 81, 551–560. simple modular architecture research tool: Identification of signaling Takahashi, T., Oie, M., and Ichihashi, Y. (1994). N-terminal amino acid domains. Proc. Natl. Acad. Sci. USA 95, 5857–5864. sequences of vaccinia virus structural proteins. Virology 202, 844– Searles, R. P., Bergquam, E. P., Axthelm, M. K., and Wong, S. W. (1999). 852. 192 LEE, ESSANI, AND SMITH

Tamin, A., Esposito, J., and Hruby, D. (1991). A single nucleotide sub- van Eijl, H., Hollinshead, M., and Smith, G. L. (2000). The vaccinia virus stitution in the 5Ј-untranslated region of the vaccinia N2L gene is A36R protein is a type Ib membrane protein present on intracellular responsible for both ␣-amanitin-resistant and temperature-sensitive but not extracellular enveloped particles. Virology 271, 26–36. phenotypes. Virology 182, 393–396. Van Meir, E., and Wittek, R. (1988). Fine structure of the vaccinia virus Tamin, A., Villarreal, E. C., Weinrich, S. L., and Hruby, D. E. (1988). gene encoding the precursor of the major core protein 4 a. Arch. Nucleotide sequence and molecular genetic analysis of the vaccinia Virol. 102, 19–27. virus HindIII N/M region encoding the genes responsible for resis- Vieira, P., de Waal-Malefyt, R., Dang, M. N., Johnson, K. E., Kastelein, R., tance to ␣-amanitin. Virology 165, 141–150. Fiorentino, D. F., deVries, J. E., Roncarolo, M. G., Mosmann, T. R., and Tartaglia, J., and Paoletti, E. (1985). Physical mapping and DNA se- Moore, K. W. (1991). Isolation and expression of human cytokine quence analysis of the rifampicin resistance locus in vaccinia virus. synthesis inhibitory factor cDNA clones: Homology to Epstein–Barr Virology 147, 394–404. virus open reading frame BCRFI. Proc. Natl. Acad. Sci. USA 88, Thompson, J. D., Higgins, D. G., and Gibson, T. J. (1994). CLUSTAL W: 1172–1176. Improving the sensitivity of progressive multiple sequence alignment Weinrich, S. L., and Hruby, D. E. (1986). A tandemly-oriented late gene through sequence weighting, position-specific gap penalties and cluster within the vaccinia virus genome. Nucleic Acids Res. 14, weight matrix choice. Nucleic Acids Res. 22, 4673–4680. 3003–3016. Tiffany, H. L., Lautens, L. L., Gao, J. L., Pease, J., Locati, M., Combadiere, Weir, J. P., and Moss, B. (1983). Nucleotide sequence of the vaccinia C., Modi, W., Bonner, T. I., and Murphy, P. M. (1997). Identification of virus thymidine kinase gene and the nature of spontaneous frame- CCR8: A human monocyte and thymus receptor for the CC chemo- shift mutations. J. Virol. 46, 530–537. kine I-309. J. Exp. Med. 186, 165–170. Whitehead, S. S., and Hruby, D. E. (1994). A transcriptionally controlled Turner, P. C., Musy, P. Y., and Moyer, R. W. (1995). Poxvirus serpins. In trans-processing assay: Putative identification of a vaccinia virus- “Viroceptors, Virokines and Related Immune Modulators Encoded by encoded proteinase which cleaves precursor protein P25K. J. Virol. DNA Viruses” (G. McFadden, Ed.), pp. 67–88. R. G. Landes, Austin. 68, 7603–7608. Upton, C., Macen, J. L., and McFadden, G. (1987). Mapping and se- Willer, D. O., McFadden, G., and Evans, D. H. (1999). The complete quencing of a gene from myxoma virus that is related to those genome sequence of Shope (rabbit) fibroma virus. Virology 264, encoding epidermal growth factor and transforming growth factor ␣. 319–343. J. Virol. 61, 1271–1275. Wolffe, E. J., Weisberg, A. S., and Moss, B. (1998). Role for the vaccinia Upton, C., Macen, J. L., Wishart, D. S., and McFadden, G. (1990a). virus A36R outer envelope protein in the formation of virus-tipped Myxoma virus and malignant rabbit fibroma virus encode a serpin- actin-containing microvilli and cell-to-cell virus spread. Virology 244, like protein important for virus virulence. Virology 179, 618–631. 20–26. Upton, C., Opgenorth, A., Traktman, P., and McFadden, G. (1990b). Xiang, Y., and Moss, B. (1999a). Identification of human and mouse Identification and DNA sequence of the Shope fibroma virus DNA homologs of the MC51L–53L-54L family of secreted glycoproteins topoisomerase gene. Virology 176, 439–447. encoded by the molluscum contagiosum poxvirus. Virology 257, Upton, C., Stuart, D., and McFadden, G. (1991). Identification and DNA 297–302. sequence of the large subunit of the capping enzyme from Shope Xiang, Y., and Moss, B. (1999b). IL-18 binding and inhibition of interferon fibroma virus. Virology 183, 773–777. gamma induction by human poxvirus-encoded proteins. Proc. Natl. Upton, C., Stuart, D. T., and McFadden, G. (1993). Identification of a Acad. Sci. USA 96, 11537–11542. poxvirus gene encoding a uracil DNA glycosylase. Proc. Natl. Acad. Xue, F., and Cooley, L. (1993). Kelch encodes a component of intercel- Sci. USA 90, 4518–4522. lular bridges in Drosophila egg chambers. Cell 72, 681–693. Vanderplasschen, A., Mathew, E., Hollinshead, M., Sim, R. B., and Yang, W. P., and Bauer, W. R. (1988). Purification and characterization of Smith, G. L. (1998). Extracellular enveloped vaccinia virus is re- vaccinia virus structural protein VP8. Virology 167, 578–584. sistant to complement because of incorporation of host comple- Yuen, L., and Moss, B. (1987). Oligonucleotide sequence signaling ment control proteins into its envelope. Proc. Natl. Acad. Sci. USA transcriptional termination of vaccinia virus early genes. Proc. Natl. 95, 7544–7549. Acad. Sci. USA 84, 6417–6421.