Copvright 0 1990 by the Genetics Society of America

Transvection in the Drosophila Ultrabithorax : A Cbx' Mutant Induces Ectopic Expression of a Normal Allele in Trans

James E. Castelli-Gair, Jose-Luis Micol'** and Antonio Garcia-Bellido Centro de Biologia Molecular, C.S.I.C., Uniuersidad Autdnoma de Madrid, 28049 Madrid, Spain Manuscript received January 20, 1990 Accepted for publication May 29, 1990

ABSTRACT In wild-type Drosophila melanogaster larvae, the Ultrabithorax (Ubx)gene is expressed in the haltere imaginal discs but not in the majority of cells of the wing imaginal discs. Ectopic expression of the Ubx gene in wing discs can be elicited by the presence of Contrabithorax (Cbx)gain-of-function of the Ubx gene or by loss-of-function mutations in Polycomb (PC)or in other trans-regulatory which behave as repressors of Ubx gene activity. Several Ubx loss-of-function alleles causethe absence of detectable Ubx proteins (UBX) or the presence of truncated UBX lacking the homeodomain. We have compared adult wing phenotypes with larval wing disc UBX patterns in genotypes involving double mutant carrying in cis one of those Ubx mutations and the Cbx' mutation. We show that such double mutant genes are (I) active in the same cells in which the single mutant Cbx' is expressed, although they are unable to yield functional proteins, and (2) able to induce ectopic expression of a normal homologous Ubx allele in a part of the cells in which the single mutant Cbx' is active. That induction is conditional upon pairing of the homologous chromosomes (the phenomenon known as transvection), and it is not mediated by UBX. Depletion of Pc gene products by Pc3 mutation strongly enhances the induction phenomenon, as shown by (I) the increase of the number of wing disc cells in which induction of the homologous- allele is detectable, and (2) the induction of not only a paired normal allele but also an unpaired one.

considerable amount of genetic, developmental during the developmentof T3 imaginal discs to main- A and molecular information is now available on taintheir proper morphogeneticidentity (MORATA genes of the bithorax complex, genes which control andGARC~A-BELLIDO 1976). Two groups of trans- the metameric identity of some thoracic and all the regulatory genes are known to be needed to maintain abdominal segments of Drosophila (LEWIS 1978; re- the correct spatial pattern of Ubx gene activity during viewedin MORATA, SANCHEZ-HERREROand CASA- development: (1) those of the Regulator of bithorax NOVA 1986;DUNCAN 1987; PEIFER, KARCH and (alsocalled trithorax) genegroup, supposed to be BENDER1987; MAHAFFEYand KAUFMAN1988; SAN- necessary for thepositive control of Ubx gene activity CHEZ-HERRERO, CASANOVAMORATA and 1988). One (INGHAM l984,1985a,b;INCH AM^^^ WHITTLE 1980; of those genes is Ultrabithorax (Ubx),which is respon- CAPDEVILAand GARC~A-BELLIDO 198 1; SHEARN sible for the segmental identities of the anterior and 1989), and (2) those of the Polycomb (PC)gene group, posterior compartmentsof the third thoracic segment supposed to be repressors (LEWIS 1978; DUNCAN and (T3a and T3p) and the anterior compartmentof the LEWIS1982; CAPDEVILA,BOTAS and GARC~A-BELLIDO first abdominal segment (Ala). The geneis also active 1986; STRUHL198 1 ; STRUHLand AKAM 1985; DUN- in some cells of the posterior compartment of the CAN 1982; SATO, RUSSELLand DENELL1983; DURA, second thoracic segment (T2p). The realm of action BROCKand SANTAMARIA 1985;JURGENS 1985). Loss- of the Ubx gene has been inferred from the pheno- of-function mutations in genes of thePC group result types ofits mutants in larval and adult cuticule(LEWIS in the ectopic expression of the Ubx gene in the T2 1978)and corroborated by meansof RNA in situ segment. hybridization(AKAM 1983; AKAM and MART~NEZ- Loss-of-function mutants of the Ubx gene include ARIAS 1985) and by immunofluorescent staining of Ubx alleles, which alter the protein coding portion of Ubx proteins (UBX) using the FP3.38 antibody, an the gene (the Ubx transcription unit, see Figure l), antibody thought to recognize all the different UBX and theabx, bx, pbxand bxd groups ofrecessive alleles. obtained by differential splicing (WHITEand WILCOX These are located within the introns of the Ubx unit 1984). orupstream the Ubx unit; they presumably affect The continuous activity of theUbx gene is required diverse regulatory regions (BENDERet al. 1983, 1985; BEACHY,HELFAND and HOGNESS1985; HOGNESSet ' Present address: Division of. Biology 156-29, (:alifornia Institute of Tecllnology. P;ls:ldetxl. Calif'ornia 91 125. al. 1985; AKAM et al. 1985; WEINZIERLet al. 1987; I' To ~11otnall corrrspondencr should be addressed. LIPSHITZ, PEATTIEand HOGNESS1987; O'CONNORet

(;rnrti<\ 126: 177-1X4 (Septcmlwt-. 1990) 178 J. E. Castelli-Gair, J. L. Micol and A. Garcia-Bellido

FIGURE1 .-Molecular map of the Ubx gene showing mutations used in thiswork. Rectangles indicate dele- tions and triangles insertions. Tran- scription units are indicated as heavy horizontal arrows, showing exons as -120 -100 -80 -60 -40 -20 0 20 kb rectangles. Thetriangle dotted is at1 I I I I I J insertion of the gypsy. . transposon,the arrow indicating the direction of its transcription. bx3 is associatedwith twoadjacent transposon insertions (gypsyand Doc), the second being irrelevant to the mutant phenotype. DfP9 Cbx ‘ I f Cbx’ corresponds to the inserrion, in I 1 invertedorientation, of theDNA pbx‘ segmentdeleted in pbx’. Df(3R)PY bx’ bxd late untt deletes the entire bithorax complex, + its left end appearing in the drawing. Ubx unlt Ubx’ corresponds to the insertion of 4 the Doc transposon in the 5’ untrans- latedregion of the first Ubx unit n I 00 exon, Ubx’”’ to a singlenucleotide 0 0 &lot changeintroducing a nonsense co- UbxP” Ubx 19’ Ubx ’ don in the exonat -53 kb, andUbx”” to ;I 1.5-kbdeletion including the honleobox. All datafor this figure wereobtained from BENDER et al. 1983, 1985; PEIFERand RENDER 1986; WEINZIERLet al. 1987. al. 1988). Gain-of-function mutants of the Ubx gene the gain-of-function Cbx’ mutation and loss-of-func- are generically termed Contrabithorax(Cbx) alleles tion Ubx mutations which prevent the production of (LEWIS1978, 1982; CASANOVA, SANCHEZ-HERRERO normal UBX. Expression was studied under different and MORATA 1985; WHITE and AKAM 1985; MICOL conditions of pairing between homologous chromo- and GARC~A-BELLIDO 1988;GONZALEZ-GAITAN, MI- somes and in the presence or absence of the PC’ allele. COL and GARC~A-BELLIDO 1990). Theyassociated are These studies were carried out in the wing disc where with ectopic expression of the gene in T2 regions in the Ubx gene is normally not expressed and where any which the wild-type allele is inactive. The best studied ectopic expression of the Ubx gene should be obvious. of those gain-of-function alleles is Cbx’ which arose as The results obtained provide evidence (1)of transvec- a transposition of a17-kb DNA segmentfrom the tion visualized at the level of protein expression, (2) upstreamregion to the second intron of the Ubx that the Cbx’ mutant allele “induces” ectopic expres- transcription unit (LEWIS 1982; BENDERet al. 1983; sion of the Ubx unit of a normal homolog, (3) that CASANOVA,SANCHEZ-HERRERO and MORATA 1985) this phenomenon is not mediated by UBX, and (4)of (Figure 1). the increased efficiency of this induction when levels Combinations of some mutant alleles of the Ubx of PC+ product are reduced. The term “induction” gene show transvection, or synapsis-dependent com- will be used throughout this paper without specific molecular implications, just to formally describe the plementation, as first described by LEWIS (1954) for activation of a Ubx’ allele by a Cbx’ alelle in trans. that gene and later observed forother Drosophila loci (reviewed in JUDD 1988; Wu and GOLDBERG1989). MATERIALS AND METHODS In transvection phenomena the phenotype of a trans heterozygote between two different alleles of a gene Fly stocks and culture: Flies were cultured on standard changesdepending upon whether the homologous medium under uncrowded conditionsat 25 f 1 O. All genetic variants used in this work have been previously described alleles are paired. Although several hypotheses have [LINDSLEYand GRELL(1968) and other references in the been proposed, little is known about the biological text]. meaning and the molecular basis of transvection ef- Immunofluorescence staining: Larvae wereobtained, fects. dissected and stained with FP3.38 antibody as described in In this paper we examine variations in the ectopic CABRERA,BOTAS and GARC~A-BELLIDO 1985. At least 20 imaginal discs of each genotype were studied. expression of the Ubx gene in the T2 segment, varia- tions revealed both by the pattern of UBX in larval RESULTS wing discs and by the morphologies of the adult wing structures. Thesevariations were studied in genotypes Adult wing phenotypes suggest that theCbx’ mu- involving double mutant chromosomes carrying in cis tant allele induces ectopic expression of a paired Transvection by Homolog Induction 179

TABLE 1

Adult wing phenotypes and UBX wingdisc patterns in genotypesused in this work

PC+ PC Adultwing UBX pattern Adultwing UBX pattern Genotypes phenotype Genotypes in wing discs phenotype in wing discs

1 +/Of(3R)P9 A a A a 2 +/+ A b a B 3 Dp(3;1)Pll5;+/Df(3R)P9 A b a B 4 Cbx’/+ D d E e 5 Cbx’Ubx’/+ C C D-E d-e 6 CbxiUbx’-zz/+d C D-E d-e 7 Dp(3;1)P115;Cbx’/Df(3R)PY D d E e 8 Dp(3;l)Pl15;CbxJUbx’/Df(3R)P9 A a C C 9 Dp(3;l)Pl15;Cbx’UbxY.ZZ/Df(3R)P9 A d C d-e 10 Dp(3;l)Pl15;Cbx’Ubx’/UbxY~22 A C C d-e 11 Dp(3;l)PI15;Cbx’Ubx’/Ub~’’~ A C C d-e 12 Ubx‘/+ A a A a 13 Ubx’.zz/+ A a A b 14 U~X“~/+ A a A b 15 Dp(3;1)P115;Ubx’/Df(3R)P9 A ND A ND 16 Dp(3;1)P115;UbxY~ZZ/Df(3R)P9 A ND A ND Wing phenotypic classes (A to E) and UBX patterns in discs (a to e) aredefined in Figure 2. ND indicates not determined. Dp(3;l)Pl15is a translocation [Tp(3;1)20;89B7-8;89B7-8]of the BX-C from 3 to the heterochromatin of the X chromosome (LINDSLEYand GRELL 1968). Df(3R)PY is a deficiency (89D9-E1;89E4-5) forthe entire BX-C (LINDSLEYand GRELL 1968). We have used the Dp(3;l)P115;Df(3R)P9combination to study genotypes in which the Ubx gene is located on another chromsome. Similar results were obtained using Tp(3;3)P146,in which the BX-C is located in the 3L chromosomal arm [Tp(3;3)64C-E;89Dl-2;9ODl](LINDSLEY and GRELL1968). It has been previously reported that the Cbx’/+ mutant wing phenotype is slightly stronger than that of Cbx’/Df(3R)P9(CASANOVA, SANCHEZ- HERREROand MORATA1985). This small difference, which can be interpreted in terms of induction of the normal chromosome by Cbx’,is not considered here, both phenotypes being classified as class D.

normal Ubx gene: Table 1 gives a summary of the type allele of Ubx translocated to anotherchromosome results obtained, on the basis of phenotypical classes (genotypes 8 and 9),a condition in which homologous defined as representedin Figure 2. The Cbx’ mutation BX-C regions are expected to be unpaired. In these causes homeotic transformations of the wing toward combinations the Cbx phenotypesare further reduced haltere, transformations which can be visualized both compared to their controls (genotype5 and 6 respec- in the wing imaginal disc and in the adultwing (Figure tively), yielding an entirely wild-type wing. This find- 2, D and d). The extentof this transformation can be ing supportsE. B. LEWIS’inference that some Cbx’Ubx modified by the presence of different cis-associated chromosomes are capable of inducing the transcrip- Ubx mutant alleles and by the degree towhich homol- tion of the Ubx homologous gene, only when both are ogous Ubx gene regions are somatically paired. We paired; but cannot do it if the Ubx wild-type allele is have employed Ubx mutant alleles which are them- in a translocated Dp(3;I)PIIS chromosome segment selves morphogenetically inactive since they either (genotypes 8 and 9). The proposed induction of the lack detectable protein products (Ubx’) or produce wild-type Ubx gene by a Cbx’Ubx chromosome is con- FP3.38-immunoreactive but truncated products lack- firmed by the wild-type phenotype of the adult wing ing the homeodomain (Ubx’.’’, Ubx’”) (WEINZIERLet when the homologous Ubx gene is also mutant and al. 1987; see Figure 1). cannot produce functional UBX (genotypes 10 and We will first consider experiments performed in a 11). PC+ geneticbackground. In cis-combinations with Visualization of UBX expression patterns in wing Cbx’, both Ubx’ and Ubx’.’’ mutations revert the Cbx discs demonstrates that the Cbx’ mutant allele in- adult wing phenotype (compare Figure 3, d with f), duces ectopic expression of a paired normal Ubx though not completely (compare genotype 4 with 5 homologous gene: Analysis of the UBX pattern of and 6 in Table 1 and Figure 2, C with D). It has been expression in the same combinations shows a major proposed for Cbx’Ubx’/+ thatthe remnant mutant difference between Cbx’Ubx’ and Cbx’Ubx’.’’ cis dou- wing phenotype is due toectopic activity of the normal ble mutants. The patternsare consistent with the Ubx homolog, induced by the Cbx’Ubx chromosome if expectation(WEINZIERL et al. 1987)that the Ubx’ pairing between homologous Ubx chromosomal re- allele does notproduce immunoreactive product while gions is not disturbed (the phenomenon designated Ubxy.22does. Thus, in genotype 8, for example, the “transvection;” LEWIS 1985).In order to test this wild-type UBX pattern reflects the behavior of the hypothesis we have studied the same genetic combi- normal, unpaired Ubx allele [Dp(3;I)PIZS],which re- nations in structural heterozygotes that have the wild mains inactive. By contrast, we infer that the aberrant 180 J. E. Castelli-Gair,J. L. Micol and A. Garcia-Bellido

a A ascertained in wing discs because the Ubx’.’’ proteins show a pattern similar to that observed in Cbx’ discs, which is superimposed to that of the normal UBX arising from the homolog (Figure 3, a and d). The Pc3 mutationenhances the induction: The presence in the genome of the Pc3 mutation in heter- b ozygotes causes a weak derepression of the wild-type Ubx gene in T2, visualized both by immunofluores- cence in wing discs and by the adult phenotype (Figure 2, B and b). This mutant phenotype (1) is dependent on the number of doses of the Ubx gene, being vir- tually wild type in the presence of only one normal c dose of Ubx but becoming stronger with increasing doses (CAPDEVILA,BOTAS and GARC~A-BELLIDO 1986; BOTAS,CABRERA and GARC~A-BELLIDO 1988) (com- pare genotypes 1 and 2 in Table I), and (2) is not dependent on the location in the genome of copies of the Ubx gene (compare genotypes 2 and 3 in Table d 1). The Polycomb adult wing phenotype disappears in all heterozygotes between the wild-type allele and a Ubx mutation, but not so the aberrant UBX wing disc pattern(compare genotype 1 with 12,13 and 14). This pattern is retained in UbxY.’’ and U~X’~~hetero- e zygotes, showing that these mutations (and possibly E Ubx’ as well) do not affect the regulatory element(s) controlled by PC+.The Pc3Cbx’ combination shows an extreme Cbx phenotype in both paired and unpaired conditions (genotypes 4 and 7). Pairing effects can be appreciated,however, in PC’ and Cbx’Ubx’ or FIGURE2.”Schemdtic representation of UBX wing disc patterns Cbx’Ubxy.22combinations (compare genotypes 5 with (a to e) and adult wing phenotypes(A to E) defining classes used in 8 and 6 with 9). Inthe former case the lack of Table 1. Shaded areas indiscs indicate positive immunofluorescence homologous pairing reduces boththe adultwing phe- labelling against UBX. Mutant transformations toward haltere are notype and imaginal disc UBX expression and in the shown in wings by the presence of haltere-like cells (shaded areas latter (as expected) only the adult wing phenotype. in B, D and E), and by the absence of wing structural elements such as the allula or some vein segments(in B and C). Theseare Interestingly in Pc3Cbx’Ubx/+ combinations both the accompanied by reductions of wing surface(as B, C, D and E show). pattern of UBX expression in the wing disc and the Although we have observed variations in phenotype among individ- adult wing phenotype aresimilar to those obtained in uals of a given genotype, classes defined in the drawing do not Cbx’/+ individuals. This indicates that the spatial pat- overlap. tern of ectopic expression of the normal Ubx allele in UBX pattern of genotype 9 reflects the superposition theseexperiments is imposed by the Cbx’ chromo- of the wild-type pattern with that of the immunoreac- some, at a low level in a PC+background and ata high tive protein products of Ubx9.” allele, derepressed by level in a Pc3 one. Individuals with genotypes 10 and Cbx’. That this pattern is quite similar to that of the 11, in which the putatively inducedchromosomes single mutant chromosome Cbx’ (genotype 7) suggests yield nonfunctional UBX, show Cbx wing disc UBX that thepresence of the Ubx’.’’ mutation in cis to Cbx’ patterns, but nearly wild-type adult wing phenotypes. does not alter the spatial specificity of that mutation. These observations support again the idea that Cbx‘ Immunofluorescence patterns confirm that Cbx’Ubx’ is imposing its own spatial pattern of expression to the is able to induce the synthesis of UBX by its normal homolog. homolog only when they are paired (compare geno- Remarkably, the enhancement by PC’ of the induc- types 5 and 8; Figure 3b). This interpretation is veri- tion phenomenon is observable even when the hom- fied in genotypes 10 and 1 1 wherethe UBX products ologs are unpaired. As seen in genotypes 8 to 11, the present in discs,although not contributingto the adult Cbx’Ubx chromosomes, which are unable to induce wing phenotype, must derivefrom the Ubx9.” and ectopic activity of the translocated Ubx gene in a PC+ Ubx19’ chromosomes. The adult wing phenotype of background, are able to do so (albeit to a low extent) genotype 6 suggests that the Cbx’Ubx9.’’ chromosome, in the presence of the PC’ mutation. This effectis not like Cbx’Ubx’, is able to induce expression of a paired, explicable as a direct effect of decreased repression normal homolog. However, this cannotbe directly by PC on the translocated Ubx gene: comparison of Transvection by Homolog Induction 181

FIGURE3.-Representative ex- amples of UBXwing disc patterns and wingphenotypes of genotypes studiedin this work. (a) Anti-UBX d immunofluorescencestaining by FP3.38 antibody of a Cbx’UbxY.22/+ wing disc, showing positive signal in mostposterior compartment cells. This UBX pattern is classified as d in Figure 2. (b)UBX pattern in a Cbx’Ubx’/+ wing disc, which corre- sponds to class c in Figure 2. This pattern is quitesimilar to that found in Dp(3;l)PI 15;Pc3Cbx’Ubx’/ e Df(3R)P9. (c) UBXpattern in a Pc’Cbx’Ubx’/+ wing disc, which cor- responds to class e in Figure 2. Signal is observed in all posterior and some anterior compartmentcells. (d) Adult wing phenotype of a Cbx’UbxY.2Z/+ individual, which corresponds class to C in Figure 2. The mutant phenotype is identical to theobserved in t Cbx’Ubx’/+ flies, despite their differ- f ent UBXwing disc patterns (com- pareFigure 3a with 3b). (e) Adult wingphenotype of a Dp(3;I)- PI 15;Pc3Cbx‘Ubx‘/Df(3R)P9 individ- ual. Themutant transformation is similar to that shown in d. (0 Adult wing phenotype of a Pc3Cbx’Ubx’/+ individual, which is classified as class D inFigure 2. Thisphenotype is similar to that of Cbx’/+ individuals. genotypes 8 to 1 1with those of 15 and 16(see column DISCUSSION of adult phenotypes in the presence of Pc3) show that Cbx’ is required to obtain an appreciable activation of A mutant allele induces in trans the ectopic activ- the Dp(3;Z)PZZ5.This is even clearer in the compar- ity of anormal homolog: Transvection hasbeen isonof genotype 3, havingtwo normal Ubx genes defined as complementation dependent on pairing derepressed by Pc3 but showing little adult mutant between homologous alleles [LEWIS(1 954); reviewed phenotype, with genotypes 8 to 11, which have only in JUDD (1988) and Wu and GOLDBERC(1989)l. In one normal Ubx gene able to contribute to themutant the vast majority of cases mutations involved in trans- phenotype. vection effects are loss-of-function allelesand comple- mentation is decreased if pairing is disrupted. Most of and Ubflz2 alleles produce normal UBX the studies of transvection effects have been largely patterns in the haltere disc: The results described inferential, given that only adult mutant phenotypes above were obtained by studying ectopic expression were considered. We show here acase in whicha gain- the gene in the T2 segment. In addition, we of Ubx of-function mutation (Cbx’) imposes aberrant behav- have studied the behavior of Ubx’, Ubxt9’ and Ubx9.” ior (ectopic expression in T2) upon a normal allele of alleles in T3, the normal realm of action of the gene, the gene located in the . in the absence of Cbx’ mutation and in a PC+back- This has been studied under conditions in which pu- ground. Ubx’ heterozygotes with strong loss-of-func- tative inducer and induced gene activitiescan be tion recessive mutations (bx3 and pbx’; see Figure 1) distinguished because (1)the mutant Cbx’ allele carries show mutant phenotypes in the adult haltere (hom- a second mutation which prevents the production of eotic transformations towards wing) which are associ- morphogenetically active UBX, and (2) the normal ated to the absenceof UBX in the corresponding Ubx allele is inactive in the tissues under study. Induc- presumptive regions of the haltere disc. Similar wing tion was inferred from the adult wing phenotype and phenotypes were observed in heterozygotes involving demonstrated by visualization of UBX in wing discs, the same recessives and Ubx9.” or Ubxt9’ mutations. thus providing evidence of transvection in the Ubx The UBX haltere disc patterns were in these cases gene at thelevel of protein synthesis, and corroborat- almost wild-type. Therefore, Ubx9.22and Ubx”’ muta- ing a previous hypothesis of E. B. LEWIS,which was tions do not alter the spatial pattern of expression of supported only by observations on mutant pheno- the Ubx gene in T3. types. 182 J. E. Castelli-Gair,J. L. Micol and A. Garcia-Bellido

The inductionphenomena studied hereare not and BRUTLAG1986). Some authors have speculated mediated by UBX since (1) the inducer chromosomes that these transcription factors could be short-radius- lack UBX (Cbx’Ubx’) or produce UBX lacking the of-action regulatory RNAs transcribed fromthe genes part of the molecule including the homeodomain involved in the transvection phenomenon (JACK and (Cbx’Ubxy.22),and (2) the induced chromosomes show JUDD1979; MICOLand GARC~A-BELLIDO1988; MICOL, the induction effect irrespectively of being able to CASTELLI-GAIRand GARC~A-BELLIDO1990; D. yield normal UBX, i.e.: the Ubxy.22and UbxIy5single MATHOG,personal communication). If Ubx RNAs of mutant chromosomes are induced in the same way in short radius of action were the inductive signals pro- which the wild-type one is. In addition, it should be duced by a Cbx’ chromosome, they would beable noted that a phenomenon dependent upon pairing ectopically to activate a paired Ubx normal gene. If between alleles located in homologous chromosomes their production were increased, or their reception can not easily be explained in terms of the protein favoured as an effect of the PC’ mutation they would products of the interacting alleles. be able to activate also the distant, nonpaired copy of Although all the induction effects presented here the Ubx gene. involve Cbx’ mutation, transvection in T2 cells is not The role of Polycomb in transvection: The inducer an exclusive property of the Cbx’ allele. Spontaneous Cbx’ chromosome has a UBX pattern in wing discs revertants of Cbx’ (CbxJRM)and Cbx2(Cbx2RM) are (see genotypes 7 and 9, in a PC+background; the known to be involved in transvection effects (MICOL pattern is essentially the same that the one shown in and GARC~A-BELLIDO1988) and one revertant of Figure 3a) which is different from that observed for Cbp’ (CbpJRM)has been obtained which, in the pres- the induced chromosome (see Figure 3b and geno- ence of PC’, is able to induce ectopic expression of a types 5, 10, and 1 1, in a PC+background): the induced normal homolog (I. E. CASTELLI-GAIR,unpublished chromosome produces detectable UBX only in a sub- results). However, on the basis of our present results set of the cells in which the inducer chromosome does it is not possible to determine if induction phenomena so. The different spatial patterns of inducer and in- involving Cbx mutationsevidence, by ectopicde- duced gene activities suggest that transvection occurs repression in T2, functions normally involved in the only in a subset of the cells where Cbx’ is expressed. regulation of the gene in T3 or if they are merely a This observation might be explained (1) by differen- consequence of the aberrant behavior of Cbx muta- tial distribution in the cells of the anlage of inductive tions. abilities of Cbx’, being gene expression and inductive Implications for transvectionmodels: There is competence noncorrelated properties, or (2) by dif- increasing evidence of regulation of transcription by ferential distribution of receptiveness to induction by interactions between proteinsbound to separateDNA the homologous allele. When the PC’ mutation is also sites (PTASHNE1986, 1988). Such actionat a distance present in the genome, transvection becomes detect- in DNA has been proposed to be possible not only able (bothin UBX disc patterns and adultphenotype) between different regionsof a gene, butalso between in all the cellsin which Cbx’ is active. Since PC+ two homologous alleles if they are paired (BENSON products aresupposed to be transcriptional repressors and PIRROTTA1988). Interactions between some ac- of Ubx gene activity, these results could beinterpreted tivated regulatory element in Cbx‘ and the promoter as an enhancement of the transcription of the induc- of the normal homolog could be taken as an expla- tive signal, arisingfrom the Cbx’Ubx chromosomes nation for the transvection effects presented in this and reaching the homologs and/or to an increase in work. However, we find difficult to explain under that the receptiveness in the induced chromosome. hypothesis the induction by Cbx’ of both the paired Our results show that PC+ products act preventing and the translocated alleles, as observed in the pres- Cbx’ dependent transvection to occur in a large num- ence of PC’ mutation (genotypes 8 to 11).Pairing ber of wing disc cells. Analogous results have been between the activated regulatory element of Cbx’ and obtained in studies of the strong mutant phenotype the two promoters should be assumed, requiring ex- of some heterozygotes involving the loss-of-function planation the differential effectiveness of the induc- pbx’ mutation, which is rescued by Pc3 (CAPDEVILA, tion effect on the promoter of the pairedallele (which BOTASand GARC~A-BELLIDO1986; J. E. CASTELLI- is induced at a lowlevel in a PC+background and GAIRand A. GARC~A-BELLIDO,manuscript in prepa- induced at a high level in a PC’ background) and on ration) and by super sex combs (a Polycomb-like muta- thepromoter of the translocated,unpaired allele tion; INGHAM 1984). It is interesting to note that the (which is not inducedin a PC+background but induced same DNA segmentabsent in the pbx’ deletion is at a low level in a PC’ background). ectopically inserted in the Ubx transcription unit in As an alternative hypothesis to explain transvection, Cbx’ mutant (Figure 1). However, the PC’ mutation some have suggested the involvement of diffusible interacts similarly with Cbd‘IRM,which is, as far as we transcriptionalfactors, with differentialdistribution know, not related to pbx’. In spite of these observa- inside the cellular nucleus (LEWIS 1985; KORNHER tions, we think it unlikely that oneof the normal roles Transvection by Homolog Induction 183 of PC+during development is to prevent transvection. Juan March. J. E. C.4. was British Council-CSIC doctorate fellow It has been shown that PCgene products repress other and J.L.M. was Fundacibn Juan March postdoctoral fellow. selector genes in cells where Ubx gene is active (WE- DEEN, HARDINCand LEVINE1986). Therefore, we LITERATURE CITED think it more likely that the inductive signal due to AKAM,M. E., 1983 The locationof Ultrabithorax transcripts in Cbx’ in T2 is associated with positive autoregulation Drosophila tissue sections. EMBO J. 2: 2075-2084. of the Ubx gene in T3. In this light, transvection AKAM,M. E., and A. MART~NEZ-ARIAS, 1985The distribution of involving Cbx alleles would be interpreted as the ac- Ultrabithorax transcripts in Drosophila embryos. EMBO J. 4: cidental activation of a normal allele by an autoregu- 1689- 1700. AKAM,M. E., A. MART~NEZ-ARIAS,R. WEINZIERLand C. D. WILDE, latory mechanism which is ectopically active inits 1985 Function and expressionof Ultrabithorax in the Dro- homolog. sophila embryo. Cold Spring Harbor Symp. Quant. Biol. 50: Implications for autoregulation: Genetic studies 195-200. led to the proposal that the BX-C genes regulate the BEACHY,A,, P. S. L. HELFANDand D. S. HOGNESS, activities of other genes (LEWIS1964; GARC~A-BEL- 1985Segmental distribution of bithorax complex proteins during Drosophila development. Nature 313: 545-55 1. LIDO 1975). On the basis of developmental and genetic BEACHY,P. A,, M. A. KRASNOW,E. R. GAVISand D. S. HOGNESS, studies, we have also inferred a role for the Ubx gene 1988 An Ultrabithorax protein binds sequences near its own products in maintaining the activity of the gene itself and the Antennapedia P 1promoters. Cell 55: 1069- 1081. (GARC~A-BELLIDOand CAPDEVILA1978; CABRERA, BENDER,W., M. AKAM,F. KARCH,P. A. BEACHY,M. PEIFER,P. SPIERER,E. B. LEWISand D. S. HOGNESS, 1983 Molecular BOTASand GARC~A-BELLIDO1985; CAPDEVILA,BOTAS genetics of the bithorax complex in Drosophilamelanogaster. and GARC~A-BELLIDO1986; BOTAS, CABRERAand Science 221: 23-29. 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