Proc. Nati. Acad. Sci. USA Vol. 91, pp. 5012-5016, May 1994 Biochemistry Specific association between the human DNA repair proteins XPA and ERCC1 (ceroderma pgmentosm/nuckotlde excision repalr/proen-proteln interactn ) LEI LI*, STEPHEN J. ELLEDGEt, CAROLYN A. PETERSON*, ELISE S. BALES*, AND RANDY J. LEGERSKI*1 *Department of Molecular Genetics, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030; and tHoward Hughes Medical Institute, Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030 Communicated by Philip C. Hanawalt, March 1, 1994 (receivedfor review December 21, 1993)

ABSTRACT Processing ofDNA damage by the nucleotide- yeast (Saccharomyces cerevisiae) reporter strain Y190, a excision repair pathway in eukaryotic cells is most likely cycloheximide-resistant derivative of Y153 (16), was used. accomplished by multiprotein complexes. However, the nature Plasmids. Plasmids containing cDNA clones of XPB ofthese complexes and the details ofthe molecular interactions (ERCC3) and CSB (ERCC6) were kindly provided by Jan between DNA repair factors are for the most part unknown. Hoeijmakers (Erasmus University, Rotterdam, The Nether- Here, we demonstrate both in vivo, using the two-hybrid lands). pMAL-XPA and pMAL-XPB were kindly provided system, and in vitro, using recombinant proteins, that the by Aziz Sancar (University ofNorth Carolina at Chapel Hill). human repair factors XPA and ERCC1 specificafly interact. In pMAL-Evi-1, pMAL-MAZ, and pMAL-paramyosin were addition, we report an initial determination of the domains in provided by Archibald Perkins (Yale University, New Ha- ERCC1 and XPA that medlate this interaction. These results ven, CT), Kenneth Marcu (State University of New York, suggest that XPA may play a role in the iocalation or ld Stony Brook), and Paul Riggs (New England Biolabs), re- ofan incision complex, composed ofERCC1 and possibly other spectively. A fragment containing ERCC1 was obtained by repair factors, onto a damaged site. PCR and cloned into the vector pCDM8 (18) and designated pCD-ERCC1. The two-hybrid vectors pASi and pACT have Two groups of mutants defective in the early steps of been described in detail elsewhere (15). pAS1 contains the nucleotide-excision repair have been identified in mammalian DNA-binding domain ofthe yeast transcription factor GAL4 cells. These are the naturally occurring cell lines obtained (residues 1-147) and pACT contains the GAL4 activation from patients with the rare disorders xeroderma pigmento- domain II (residues 768-881). pACTII is a derivative of sum, Cockayne syndrome, and trichothiodystrophy and the pACT containing a multiple cloning site that facilitates the laboratory-derived rodent cell lines designated ERCC (exci- subcloning of inserts from pASi clones. Construction of sion repair cross-complementing). Eleven genetic comple- clones containing in-frame fusions in pAS1 and pACTII mentation groups have been identified in the former series (1, vectors was accomplished by standard procedures. 2) and 11 in the latter (3, 4). Several of the mutants in these Two-Hybrid Assays. Two-hybrid screening of a human two classes fall into the same complementation group; how- lymphocyte cDNA library in pACT vector, P-galactosidase ever, the full extent of overlap remains to be determined. assay, and rescue ofplasmids from yeast clones into bacterial Seven of these genes have now been cloned (5-11), setting hosts were accomplished essentially as described (15). the stage forinvestigations into how DNA damage processing Sequence Analysis. Manual sequencing ofrescued plasmids is mediated by this system. One aspect of these studies is to was accomplished by the dideoxy method using Sequenase determine the nature ofprotein-protein interactions between 2.0 according to the manufacturer's (United States Biochem- DNA-repair factors and to define the composition of multi- ical) specifications. Homology searches were performed with factorial complexes with the eventual goal ofelucidating their FASTA (Genetics Computer Group, Madison, WI). function. Deletion Mutants. Deletion mutants expressing truncated The yeast two-hybrid system developed by Fields and forms of ERCC1 and XPA were constructed by the exonu- colleagues (12, 13) is a novel genetic approach that is used for clease III method (19). Plasmid CD-ERCC1 was prepared for the detection of interacting proteins. In this system, associ- exonuclease III digestion by cleavage with the enzymes Bgl ation in yeast between two proteins, one fused to the DNA- II and Apa I. Translation termination was achieved by the binding domain and the second to the activation domain II of presence of naturally occurring stop codons in all reading GALA, activates promoters containing GALA-binding sites frames downstream of the Apa I site. For pMAL-XPA an (14). We have used a modification (15, 16) of the original oligonucleotide adaptor containing a Kpn I site and stop system involving a dual selection scheme for HIS3 and LacZ codons in all three reading frames was inserted between the to screen a human cDNA library for genes whose expression Xba I and Sal I sites. The resulting construction was cleaved products interact with the human repairfactor ERCC1 (5, 17). by Kpn I and Xba I for subsequent digestion by exonuclease In addition to the isolation of a number of novel genes, this III. The construct expressing XPA-(1-114) was prepared by screening also resulted in the identification of the repair deleting the 0.5-kb BsaBI-Xba I fiagment from pMAL-XPA. protein XPA as a factor that associates with ERCC1. The pMAL construct expressing XPA-(59-273) was kindly provided by Aziz Sancar. Preparation of Recombinant Proteins and in Vitro Binding MATERIALS AND METHODS Asay. Maltose-binding protein (MBP) fusion proteins (from Bacterial and Yeast Strains. Escherichia coli strain MC1061 pMAL constructs) were expressed in E. coli MC1061. Cul- was used for subcloning of cDNA inserts and for rescue of tures were grown to an OD600 of 0.6 and induction of fusion plasmids from yeast cells. For two-hybrid screenings the proteins was initiated by the addition of isopropyl /-D- thiogalactopyranoside to a final concentration of0.3 mM. All The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" Abbreviation: MBP, maltose-binding protein. in accordance with 18 U.S.C. §1734 solely to indicate this fact. *To whom reprint requests should be addressed. 5012 Downloaded by guest on September 27, 2021 Biochemistry: Li et al. Proc. Natl. Acad. Sci. USA 91 (1994) 5013 subsequent operations were performed at 40C. After 2 hr of Table 1. Two-hybrid analysis of ERCC1 and XPA incubation cells were pelleted and resuspended in 50 ml (per pACT liter of culture medium) of MBP buffer (20 mM Tris Cl, pH 7.4/200 mM NaCl/1 mM EDTA/1 mM dithiothreitol) plus 0.2 pASi ERCCI XPA mM phenylmethanesulfonyl fluoride and leupeptin, pepsta- ERCCJ - + tin, and aprotinin each at 0.5 jg/ml. The cell suspension was XPA + - sonicated to near clarity, centrifuged at 27,000 x g for 10 min, XPB - - and aliquots of the supernatant were stored at -80oC. For CSB - - protein examination, 20 Ad of amylose-agarose beads (New SNFI - - England Biolabs) was incubated with 800 p4 of bacterial tat - - lysate for 30 min with rocking. The beads were centrifuged at p53 gene - - 735 x g and washed four times with MBP buffer. Bound Lamin gene - - eluted with MBP buffer 10 mM maltose proteins were plus Each pASi construct contains the sequence encoding the DNA- and subsequently examined by SDS/PAGE. For binding binding domain of the yeast transcription factor GAL4 (residues assays, beads were washed twice with binding buffer [40 mM 1-147) fused in frame to each listed gene. Each pACT construct Hepes, pH 7.9/2% (vol/vol) glycerol/50 mM KCI/50 mM contains the sequence encoding the GAL4 activation domain II NaCI/5 mM MgCl2/1 mM EDTA/0.5 mM dithiothreitol/0.2 (residues 768-881) fused in frame to each listed gene. Plus sign mM phenylmethanesulfonyl fluoride with leupeptin, pepsta- indicates activation ofthe HIS3 and lacZ reporter genes in the yeast tin, and aprotinin each at 0.5 pg/mll and resuspended in 600 strain Y190. A. of binding buffer. [35S]Methionine-labeled ERCC1 was prepared with 0.5 pg of the construct pCD-ERCC1 or its MBP fusion proteins (Fig. la), which included XPA (22), deletion mutants by a coupled in vitro transcription/ XPB, and two zinc finger proteins, MAZ (23) and Evi-l (24), translation system in a total volume of 50 yl (TnT, Promega). were prepared and examined as affinity reagents for ERCC1 Ten microliters of [35S]methionine-labeled ERCC1 was when bound to amylose resin. Maz and Evi-l were chosen as added to the beads and the suspension was incubated with controls because XPA contains a zinc finger motif (7). rocking at 40C for 1 hr. After four washes with binding buffer, Following washing, bound MBP fusion proteins and any the bound proteins were eluted in binding buffer containing associated proteins were eluted with maltose and examined 10 mM maltose and examined by SDS/PAGE followed by by SDS/PAGE (Fig. lb). Only the MBP-XPA fusion protein autoradiography. bound [35S]methionine-labeled ERCC1, thus indicating a Immunoblon. HeLa whole cell extracts were prepared high-affinity interaction between ERCC1 and XPA. as described (20). Polyclonal anti-ERCC1 antiserum (21) was As a further confirmation ofthis in vitro interaction, HeLa provided by Jan Hoeijmakers. Immunoblotting was per- whole cell extracts were incubated with beads containing formed with an ECL kit (Amersham) according to the man- bound MBP-XPA fusion protein or MBP. After washing, ufacturer's specifications. bound fractions were examined by SDS/PAGE and immu- noblotting (Fig. 2). Only the fraction bound to the MBP-XPA fusion protein contained ERCC1, indicating that XPA and RESULTS ERtCC1 also interact in the context of a whole cell extract. Association of XPA and ERCC1 in Vivo. The two-hybrid Identicon of XPA and ERCC1 Interactive Domain. To system was used to screen over 2 million yeast transformants localize the regions in XPA and ERCC1 that mediate the for factors that interact with the repair protein ERCC1. Ten clones were isolated from a human cDNA library that were a 'Da 445 5 both His+ and LacZ+. To eliminate clones whose interaction 204 - was not specific for ERCC1, each rescued library plasmid was tested in the presence of pAS1 plasmids fused to the genes for Tat, p53, SNF1, and lamin for activation of HIS3 116 - and lacZreporter genes. All 10 clones were His3- and LacZ- 97 - in the presence of these constructs in the yeast reporter 67 - strain, suggesting that the interactions were specific to ERCC1. Partial sequencing of the cDNA inserts indicated 45 - that 9 contained unique sequences not present in computer data bases (GenBank and EMBL, February 1994) and hence b ; 3 4 : presumably represented unidentified human genes. The par- tial sequence of1 clone, however, had complete identity with a portion of the cDNA sequence of the repair gene XPA (7). To confirm an interaction between ERCC1 and XPA, an ERCC1 - - XPA cDNA clone was obtained from a separate source and the two-hybrid system was used to directly assay for inter- action of XPA with ERCC1. As Table 1 shows, the assay again indicated an interaction between these two repair proteins. Exchanging the cDNA inserts by placing XPA into FIG. 1. Association between ERCC1 and XPA proteins in vitro. the pAS1 vector and ERCCI into the pACTII vector also (a) SDS/PAGE analysis of purified MBP fusion proteins tested as yielded a positive result. As is also shown, we did not detect binding substrates for ERCC1: MBP-XPA (lane 1), MBP (lane 2), an interaction of either XPA or ERCC1 in the presence of a MBP-XPB (lane 3), MBP-Evi-1 (lane 4), MBP-MAZ (lane 5), and number ofcontrol fusion proteins, which included the repair MBP-paramyosin (lane 6). Molecular size markers migrated as XPB and CSB This also indicated that indicated. (b) Interaction ofP5S]methionine-labeled ERCC1 (lane 1) proteins (8) (9). assay with MBPfusion proteins. MBPfusion proteins were immobilized on neither ERCC1 nor XPA forms a homodimer. amylose-agarose beads and incubated with P5Slmethionine-labeled Association of XPA and ERCC1 in Vito. To directly dem- ERCC1. After washing, bound proteins were eluted with maltose, onstrate an interaction in vitro between ERCC1 and XPA, resolved by SDS/PAGE, and analyzed by autoradiography: MBP- [35S]methionine-labeled ERCC1 was prepared by expression XPA (lane 2), MBP (lane 3), MBP-XPB (lane4), MBP-Evi-1 (lane 5), in a coupled transcription/translation system. A series of MBP-MAZ (lane 6), and MBP-paramyosin (lane 7). Downloaded by guest on September 27, 2021 5014 Biochemistry: Li et al. Proc. Nati. Acad. Sci. USA 91 (1994) 1 2 MBP-XPA. As shown (Fig. 3b), ERCC1-(1-119) was suffi- cient for interaction with MBP-XPA, but ERCC1-(1-91) failed to bind to MBP-XPA. These results indicate that the amino-terminal 40% of ERCC1 is sufficient for binding of XPA and suggest that amino acids within residues 91-119 of ERCC1 are required for the interaction. It is important to note that these experiments do not exclude the possibility that a second XPA interactive domain exists in the carboxyl- ERCCIl- terminal portion of ERCC1. Among deletion mutants of MBP-XPA (Fig. 3d) a con- struct containing XPA residues 1-114 bound full-length ERCC1, whereas a construct containing XPA residues 1-74 failed to form a complex with ERCC1 (Fig. 3e). An amino- terminal deletion mutant lacking the first 58 residues ofXPA also bound ERCC1 (Fig. 3e). This particular mutant has been shown previously to completely complement the repair de- ficiency of xeroderma complementation group A cells (7). FiG. 2. Retention of ERCC1 by MBP-XPA in HeLa whole cell These results indicate that the amino-terminal 42% ofXPA is extracts. Binding of ERCC1 to MBP (lane 1) or MBP-XPA (lane 2) sufficient for binding of ERCC1 and suggest that residues was tested as described in the legend to Fig. 1. ERCC1 was detected 58-114 are necessary and likely to be sufficient for the by SDS/PAGE followed by immunoblotting. interaction. As indicated above, the existence of a second ERCC1 interactive domain is not excluded by these experi- identified interaction, we used the exonuclease Ill method ments. (19) to generate a series ofdeletion mutants ofboth XPA and ERCCI that expressed carboxyl-terminally truncated poly- peptides (Fig. 3 a and c). Each mutant was then examined, by DISCUSSION the in vitro method described above, for the appropriate The results of the experiments to identify the domains that interaction with the full-length form of either ERCC1 or mediate the interaction between ERCC1 and XPA are

a kDa 1 2 3 4 5 6 7 8 d WDa f... C~- - _ L.3 -w i

28 - *5-0" tS5 _- 400V 18 - ...... 5:N

.0 ,v-

b kDa 1 2 3 L. 5 6 - 8 e *3 *

28

c f ERCC1 X P.:. ]c +

1233 1,' + l:f + r + I_--I 553 - 1 5~ I 52 O~ ~~~~~~~~~

= 9 , FIG. 3. Mapping of the interactive domains in ERCC1 and XPA. (a) Autoradiograph of SDS/PAGE analysis of [35S]methionine-labeled ERCC1 deletion mutants: ERCC1 (lane 1), ERCC141-270) (lane 2), ERCC1-(1-233) (lane 3), ERCC1-(1-177) (lane 4), ERCC1-(1-153) (lane 5), ERCC1-(1-119) (lane 6), ERCC1-(1-52) (lane 7), and ERCC1-(1-91) (lane 8). (b) ERCC1 deletion mutants were tested for interaction with MBP-XPA as described in the legend to Fig. 1. Lane designations are the same as in a. (c) Schematic illustration of ERCC1 deletion mutants and the results of their interaction with MBP-XPA. Numbers at left refer to lane numbers in b. Plus or minus sign indicates interaction or no interaction with MBP-XPA, respectively. (d) Purified MBP-XPA deletion mutants were examined by SDS/PAGE followed by Coomassie blue staining: XPA (lane 1), XPA-(59-273) (lane 2), XPA-(1-237) (lane 3), XPA-(1-219) (lane 4), XPA-(1-153) (lane 5), XPA-(1-114) (lane 6), and XPA41-74) (lane 7). (e) MBP-XPA deletion mutants were tested for interaction with [35S]methionine-labeled ERCC1 as described above. Lane designations are the same as in d. Molecular size markers migrated as indicated in a and c. (f) Schematic illustration of XPA deletion mutants and the results of their interaction with ERCC1. Downloaded by guest on September 27, 2021 Biochemistry: Li et A Proc. Natl. Acad. Sci. USA 91 (1994) 5015 XPAIR HTH Radl IR 92-I -- 297 92 119

E-cluster ZF r , C I XPA I 1 273 75ERCC1 IR1 FIG. 4. Schematic of identified motifs in ERCC1 and XPA. Thick dark lines indicate regions of conservation between ERCC1 and RAD10 (5) and between XPA and RAD14 (25). Boxes indicate identified structural features. Dashed box indicates that placement ofthis feature is based upon the homology of this region with the region required for RAD1 association mapped in RAD10 (24). Numbers below boxes refer to amino acid residues that have been shown to be required but not necessarily sufficient for the indicated interaction. IR, interactive region; HTH, helix-turn-helix; E-cluster, acidic region containing seven consecutive glutamic residues; ZF, zinc finger motif. summarized in Fig. 4 along with other identified features of mented, extracts from both ERCC1 and ERCC4 mutants in these proteins. Previous studies have demonstrated that the vitro. These findings support the functional relevance of the yeast DNA repair factors RAD1 and RAD10, the yeast XPA-ERCC1 interaction by indicating that the ERCC1- homologue of ERCC1 (5), associate with each other (25, ERCC4 complex associates with XPA. 26). In Fig. 4 we have indicated the of region greatest We thank Jan Hoebmakers, Aziz Sancar, Archibald Perkins, conservation between RAD10 and ERCC1 and a region Kenneth Marcu, and Paul Riggs for generously providing reagents. previously found to be required for association of RAD10 This work was supported by National Institutes of Health Grants with RAD1 (26). Our results indicate that the amino- CA52461 (to R.J.L.) and AG11085 (to S.J.E.). S.J.E. is a Pew terminal 40%o of ERCC1 is sufficient for binding ofXPA and Scholar in the Biomedical Sciences and an Investigator of the place a required region of the XPA interactive domain near Howard Hughes Medical Institute. the amino-terminal end of the RAD10/ERCC1 conserved 1. Hoejmakers, J. H. J. & Bootsma, D. (1990) Cancer Cells 2, domain. It is interesting that the helix-turn-helix DNA- 311-320. binding motif in ERCC1 (5) is located between an XPA 2. Stefanini, M., Vermeulen, W., Weeda, G., Giliani, S., Nardo, interactive region that we have identified and a domain T., Mezzina, M., Sarasin, A., Harper, J. I., Arlett, C. F., required for RAD1 interaction (26). Hoeimakers, J. H. J. & Lehmann, A. R. (1993) Am. J. Hum. The yeast homologue ofXPA is RAD14 (27), and a region Genet. 53, 817-821. of high conservation between these two proteins is also 3. Collins, A. R. (1993) Mutat. Res. 293, 94-118. shown in Fig. 4. Our observations with deletion mutants of 4. Riboni, R., Botta, E., Stefanini, M., Numata, M. & Yasui, A. XPA indicated that XPA-(59-114) is sufficient for associ- (1992) Cancer Res. 52, 6690-6691. ation with ERCC1 and showed that the zinc finger motif 5. van Duin, M., de Wit, J., Odijk, H., Westerveld, A., Yasui, A., (residues 105-130) is not required for binding ERCC1. Koken, M. H. M., Hoeijmakers, J. H. J. & Bootsma, D. (1986) Residues 75-114 were found to contain a region that is most Cell 44, 913-923. 6. Weber, C. A., Salazar, E. P., Stewart, S. A. & Thompson, likely required for binding ERCC1. These results also L. H. (1990) EMBO J. 9, 1437-1447. indicate that the ERCC1 interactive domain lies outside of 7. Tanaka, K., Niura, N., Satokata, I., Miyamoto, I., Yoshida, the highly conserved region between XPA and RAD14. In M. C., Satoh, Y., Kondo, S., Yasui, A., Okayama, H. & fact, recent experiments have shown that the yeast homo- Okada, Y. (1990) Nature (London) 348, 73-76. logues of XPA and ERCC1 do not interact in a two-hybrid 8. Weeda, G., van Ham, R. C. A., Vermeulen, W., Bootsma, D., assay (E. Friedberg, personal communication). This result van der Eb, A. J. & Hoeijmakers, J. H. J. (1990) Cell 62, may be due to a failure of the two-hybrid system to detect 777-791. the interaction or it may indicate a mechanistic difference 9. Troelstra, C., van Gool, A., de Wit, J., Vermeulen, W., in this part of the nucleotide-excision repair pathway be- Bootsma, D. & Hoejmakers, J. H. J. (1992) Cell 71, 939-953. tween yeast and mammalian cells. 10. Legerski, R. J. & Peterson, C. A. (1992) Nature (London) 359, In it and 70-73. yeast is known that RAD10 RAD1 form a complex 11. Scherly, D., Nouspikel, T., Corlet, J., Ucla, C., Bairoch, A. & that constitutes a single-stranded DNA endonuclease (28, Clarkson, S. G. (1993) Nature (London) 363, 182-185. 29). It has been suggested that this association, most likely in 12. Fields, S. & Song, 0. (1989) Nature (London) 340, 245-246. conjunction with other repair factors, forms a damage- 13. Chien, C., Bartel, P. L., Sternglanz, R. & Fields, S. (1991) specific DNA-incision complex. In vitro studies with mam- Proc. Natl. Acad. Sci. USA 88, 9578-9582. malian cell extracts have suggested that ERCC1 forms a 14. Ma, J. & Ptashne, M. (1988) Cell 55, 443-446. highly stable complex with the complementing activities of 15. Durfee, T., Becherer, K., Chen, P.-L., Yeh, S.-H., Yang, Y., ERCC4, ERCC11, and XPF (21, 30, 31). It is likely that one Kilburn, A. E., Lee, W.-H. & Elledge, S. J. (1993) Genes Dev. ofthese latter factors is the mammalian homologue ofRAD1 7, 555-569. and that this contains an 16. Harper, J. W., Adami., G. R., Wei, N., Keyomarsi, K. & complex endonuclease that incises Elledge, S. J. (1993) Cell 75, 805-816. DNA at the site of damage. 17. Westerveld, A., Hoeijmakers, J. H. J., van Duin, M., de Wit, XPA is a zinc finger DNA-binding protein that has been J., Odjjk, H., Pastink, A. & Bootsma, D. (1984) Nature reported to possess a higher affinity for damaged double- (London) 310, 425-429. stranded DNA than for undamaged double-stranded DNA 18. Seed, B. (1987) Nature (London) 329, 840-842. (32, 33). Thus, one possible function of XPA, suggested by 19. Henikoff, S. (1987) Methods Enzymol. 155, 156-165. our results, is the loading and possible orientation of the 20. Wood, R., Robins, P. & Lindahl, T. (1988) Cell 53, 97-106. incision complex, containing ERCC1 and the factors men- 21. van Vuuren, A. J., Appeldoorn, E., Odijk, H., Yasui, A., tioned above, to the site of damage. This model would also Jaspers, N. G. J., Bootsma, D. & Hoeijmakers, J. H. J. (1993) that XPA would most with EMBO J. 12, 3693-3701. predict likely interact components 22. Park, C.-H. & Sancar, A. (1993) Nucleic Acids Res. 21, ofthe recognition and/or site-preparation complex in order to 5110-5116. effect localization of the incision complex. This model is 23. Bossone, S. A., Asselin, C., Patel, A. J. & Marcu, K. B. (1992) supported by the results of Park and Sancar (34) (the paper Proc. Natl. Acad. Sci. USA 89,-7452-7456. immediately following this one) in which a MBP-XPA fusion 24. Perkins, A. S., Fishel, R., Jenkins, N. A. & Copeland, N. G. protein was used to affinity-purify a complex that comple- (1991) Mol. Cell. Biol. 11, 2665-2674. Downloaded by guest on September 27, 2021 5016 Biochemistry: Li et al. Proc. Nat!. Acad. Sci. USA 91 (1994) 25. Bailly, V., Sommers, C. H., Sung, P., Prakash, L. & Prkash, 30. Biggerstaff, M., Szymkowski, D. E. & Wood, R. D. (1993) S. (1992) Proc. Natl. Acad. Sci. USA 89, 8273-8277. EMBO J. 12, 3685-3692. 26. Bardwell, L., Cooper, A. J. & Friedberg, E. C. (1992) Mol. 31. Reardon, J. T., Thompson, L. H. & Sancar, A. (1993) Cold Cell. Biol. 12, 3041-3049. Spring Harbor Symp. Quant. Biol. 58, 605-617. 27. Bankmann, M., Prakash, L. & Prakash, S. (1992) Nature 32. Robins, P., Jones, C. J., Biggerstaff, M., Lindahl, T. & Wood, (London) 355, 555-558. R. D. (1991) EMBO J. 10, 3913-3921. 28. Tomkinson, A. E., Bardwell, A. J., Bardwell, L., Tappe, N. J. 33. Jones, C. J. & Wood, R. D. (1993) Biochemistry 32, 12096- & Friedberg, E. C. (1993) Nature (London) 362, 860-862. 12104. 29. Sung, P., Reynolds, P., Prakash, L. & Prakash, S. (1993) J. 34. Park, C.-H. & Sancar, A. (1994) Proc. Natl. Acad. Sci. USA 91, Biol. Chem. 268, 26391-26399. 5017-5021. Downloaded by guest on September 27, 2021