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What the papers say Masters change, slaves remain Patricia Graham,* Jill K. M. Penn, and Paul Schedl Summary Sex determination offers an opportunity to address many classic questions of developmental biology. In addition, because sex determination evolves rapidly, it offers an opportunity to investigate the evolution of genetic hierarchies. Sex determination in Drosophila melanogaster is controlled by the master regulatory gene, Sex lethal (Sxl ). DmSxl controls the alternative splicing of a downstream gene, transformer (tra), which acts with tra2 to control alternative splicing of doublesex (dsx). DmSxl also controls its own splicing, creating an autoregulatory feedback loop that ensures expression of Sxl in females, but not males. A recent paper(1) has shown that in the dipteran Ceratitis capitata later (downstream) steps in the regulatory hierarchy are conserved, while earlier (upstream) steps are not. Cctra is regulated by alternative splicing and apparently controls the alternative splicing of Ccdsx. However, Cctra is not regulated by CcSxl. Instead it appears to autoregulate in a manner similar to the autoregulation seen with DmSxl. BioEssays 25:1–4, 2003. ß 2002 Wiley Periodicals, Inc. Introduction The choice of sexual cell fate is a developmental decision akin to the choice between becoming an epidermal cell or a neuron. Thus sex determination offers a chance to investigate the classical problems of developmental biology. How are choices between possible cell fates made, maintained and implemented? Studies of sex determination in Drosophila melanogaster have already made significant contributions to our understanding in this area. For example, analysis of the master switch gene, Sex-lethal (Sxl ), has shown sexual identity is remembered during the life cycle through a mechanism involving alternative splicing and autoregulation.(2,3) Examination of sex determination in different species should add to our knowledge of some of these topics and may make novel contributions to others. Sex determination shares some general features in all species. An initial signal governing the choice of sex is transmitted through a regulatory cascade to activate the genes ultimately required to produce the particular physiological and behavioral phenotypes of the two sexes. In different species Dept. Molecular Biology, Princeton University. *Correspondence to: Patricia Graham, Dept. Molecular Biology, Princeton University. NJ 08544-1041. E-mail: [email protected] DOI 10.1002/bies.10207 Published online in Wiley InterScience (www.interscience.wiley.com). BioEssays 25:1–4, ß 2002 Wiley Periodicals, Inc. the initial signals at the top of the regulatory hierarchy vary considerably. For example, vertebrates use a male determining factor on the Y chromosome.(4 –6) Two well-characterized model systems, Drosophila melanogaster and Caenorhabditis elegans use the ratio of X chromosomes to autosomes (reviewed in Ref. 7). Yet other species use autosomal factors or environmental cues. Studying sex determination in different but related species will not only add to our understanding of decision making in development, but will also help elucidate how regulatory hierarchies have evolved. Evidence from studies comparing sex determination in different species of dipteran insects reveals that the master genes at the top of the regulatory hierarchy can change dramatically as new species and genus evolve, while the slave genes at the bottom of the hierarchy remain the same, carrying out essentially identical functions from one species to the next. In this review, we discuss some of what is known about sex determination in Drosophila melanogaster and how this relates to what has recently been learned about sex determination in Ceratitis capitata. Choice and memory in Drosophila melanogaster The initial signal and the mechanisms by which that signal are translated into the female or male fate are well understood in Drosophila melanogaster (Fig. 1A, for a recent review see Ref. 8). The Sxl gene is turned on early in development in females, while it remains off in males. The presence of Sxl protein induces the productive splicing of mRNAs from the downstream target gene transformer (tra). Tra protein expressed from the female tra mRNAs acts together with Tra2 (which is expressed in both sexes) to activate the female splicing of mRNAs from the gene doublesex (dsx).(9) The Dsxf protein represses the transcription of genes required for male development and activates those required for female development. When Sxl protein is absent, tra mRNA is spliced in a nonproductive pattern, which includes exon sequences containing a stop codon. No Tra protein is expressed from the male tra mRNA and as a consequence dsx mRNA is spliced in the male pattern. The Dsxm protein translated from this mRNA represses female development and promotes male. The choice of sexual identity early in development depends upon a system that assesses the X chromosome to autosome (X/A) ratio. The target for the X/A counting system is a special Sxl establishment promoter Sxl-Pe. This promoter is turned on for a brief period in pre-cellular blastoderm female (2X/2A) BioEssays 25.1 1 What the papers say Figure 1. Sex determination compared in Drosophila melanogaster and Ceratitis capitata. The Ceratitis tra gene has the role of ‘‘master regulator,’’ taking over the position held by Sxl in Drosophila. embryos, while it remains off in male (1X/2A) embryos. The function of the Sxl proteins encoded by the Sxl-Pe mRNAs is to activate the female-specific splicing of mRNAs expressed from the Sxl maintenance promoter, Sxl-Pm. Sxl-Pm turns on in both sexes at cellularization, just as Sxl-Pe is being shut off in female embryos, and then remains on during the rest of the life cycle. The female Sxl-Pe mRNAs lack the 3rd or male exon, which contains multiple in-frame stop codons, and they encode functional proteins that have two RRM-type RNAbinding domains. These Sxl proteins, in turn, direct the female splicing of Sxl-Pm mRNAs, setting up a positive autoregulatory feedback loop which ensures the continued production of Sxl protein and thus memory of female identity. In males where Sxl-Pe is never turned on, transcripts from Sxl-Pm are spliced to include the translation terminating male exon and thus male identity is set by default. Choice and memory in Ceratitis capitata The Sxl homologue has been isolated from several dipterans including Ceratitis capitata(10–13) and is generally well conserved. Within the genus Drosophila, Sex-lethal is produced in a sex-specific manner, which is consistent with the idea that it retains its role as a master regulatory gene.(10) However, in virtually every dipteran examined outside the genus Drosophila, Sex-lethal is not expressed in a sex-specific manner, and has no apparent role in sex determination.(11,12) In fact, although Sxl is present in Ceratitis, and has 69% identity and 79% similarity to DmSxl at the amino acid level, it is no longer the master regulator of sexual identity. In this case, how is sex determined in Ceratitis and what gene(s) controls and maintains sexual identity? In Ceratitis capitata, the X/A ratio is not used to determine sexual fate. Instead, the presence of a Y-linked factor, M, 2 BioEssays 25.1 promotes male development while its absence leads to female development(14) (Fig. 1B). Currently the M factor has not been identified, and it is unknown how it promotes maleness. A recent paper(1) has shown that the Ceratitis homologue of Dmtra is functionally conserved even though it has only 32–40% identity at the amino acid level. The first evidence of functional conservation was the finding of sex-specific transcripts of Cctra. Furthermore, as in Drosophila, male-specific tra transcripts contain multiple stop codons while the female transcript encodes a SR-rich protein. To show that Cctra is essential for female development, Pane et al. (2002) inhibited Cctra function by injecting embryos with Cctra dsRNA, causing the endogenous Cctra mRNA to be degraded. The injected XX embryos developed into sexually transformed adult males while the development of the XY embryos was unaffected. Thus, despite the very poor sequence conservation, Cctra and Dmtra are functionally similar. Since the sexually transformed XX Ceratitis flies produced predominantly male dsx RNA, it would appear that Cctra controls sexual differentiation by regulating Ccdsx. Sequence analysis of Ccdsx genomic DNA and cDNAs indicates that there are Tra/Tra2-binding sites in positions similar to those in Dmdsx, and that the Ccdsx mRNAs are structurally very similar to those of Dmdsx.(12) Although it remains to be seen whether there is a tra2 homologue in Ceratitis, this suggests that the mechanism controlling the alternative splicing of dsx mRNA may also be similar in the two species. The dsRNA injections into XX embryos would be expected to result in a transient loss of tra activity. Yet the adult animals that develop from the embryos are transformed XX males and RT-PCR analysis of the tra mRNAs from these animals reveals that the message is spliced in the male pattern. One way to explain how a transient loss of tra activity is sufficient to block What the papers say the female mode of tra splicing later in development, is that Tra has a positive autoregulatory function and is required to direct the female splicing of tra mRNA. In this case, a transient loss of tra activity in XX embryos would be sufficient to induce a permanent switch in sexual identity. Consistent with this idea, examination of the Cctra gene revealed several putative Tra/Tra2-binding sites in and around the male-specific exons. If this model is correct, the role of Cctra in Ceratitis capitata would be similar to that of Sxl in the Drosophilids: it would be at the top of the sex determination hierarchy, directing female sexual development by regulating downstream target genes, and functioning to maintain female identity through its autoregulatory activity. Although the initial signal and master regulatory genes vary in different species, the target of these switch genes, the dsx gene, appears to have both structural and functional conservation across a wide range of species. The dsx gene of Megaselia scalaris shows both conserved structure and sexspecific splicing patterns.(15) dsx may also be structurally and functionally conserved in Ceratitis capitata. Intriguingly, this conservation may extend beyond dipterans as the dsx homologue in Caenorhabditis elegans, male abnormal 3 (mab-3), also controls sexual cell fate.(16,17) The role of Tra at the molecular level Previous data indicate that SR-proteins such as Tra and Tra2 generally act to activate splice sites that would otherwise not be used (reviewed in Ref. 18). This has been shown to be the case for Drosophila Tra and Tra2 for the sex-specific alternative splicing of dsx.(19) and another gene required for mating behavior, fruitless (fru).(20) In Ceratitis, as mentioned above, it has been proposed that Tra and Tra2 regulate dsx in this manner also. Surprisingly, it appears that Tra and Tra2 may use a different mechanism to regulate the splicing of Cctra mRNA. Based on the location of the putative Tra/Tra2-binding sites in the Cctra transcript and the sequences of the sexspecifically alternatively spliced Cctra isoforms, Pane et al.(1) have hypothesized that Tra and Tra2 block the use of the malespecific splice sites rather than activate a weaker splice site. Although this is a new role for Tra and Tra2, it should be noted that the context of a binding site can influence the activity of splicing regulators. For example, in adenovirus, it has been shown that SR proteins bind within an intron of the L1 transcriptional unit and are able to inhibit splicing by preventing the recruitment of the U2 snRNP to the spliceosome.(21) When the binding site is instead placed within an exon, the SR proteins are able to activate the upstream splice site. Beyond sex determination Developmental decisions are often connected to one another. Pathways regulating cell division ‘‘talk’’ with those regulating differentiation to produce the correct number of the right type of cells. Thus it is of interest to learn how related regulatory hierarchies might be connected to one another. For example, the process of sex determination is sometimes connected to a related process called dosage compensation. In many species, the two sexes contain different numbers of sex chromosomes (as in humans where the male has one and the female has two X chromosomes). Therefore expression of genes on the sex chromosomes may need to be regulated to compensate for the difference in dose between the sexes. This compensatory process is called dosage compensation. In Drosophila melanogaster, the processes of sex determination and dosage compensation are coupled by the multifunctional, master regulator Sxl. By contrast, in mammals, dosage compensation is dependent upon Xist, a noncoding RNA, while sex determination is controlled by Sry. In Drosophila, Sxl regulates dosage compensation in two ways. First, in females DmSxl represses the translation of a gene called male-specific lethal-2 (msl-2) which is required to hyperactivate expression of X-linked genes in males.(22–24) Second, DmSxl directly regulates expression of some X-linked genes.(25,26) Because the processes of sex determination and dosage compensation are coupled in Drosophila, changes at the top of the sex determination hierarchy also cause sexspecific lethality rather than just sexual transformation.(8) One side effect of this coupling might be that changes in the genes at the top of the sex determination hierarchy are more difficult, and thus the coupling might promote the evolutionary stability of the hierarchy. Dosage compensation in Ceratitis capitata has not been formally investigated. However, the paucity of X-linked mutations and the observation that the Ceratitis X chromosome is predominantly heterochromatic suggest that there may be relatively few genes on the Ceratitis X chromosome. Thus there may be little need for dosage compensation in this species. This would not be a unique situation as Musca domestica may have used the same solution to the problem of a two-fold difference in gene dose between the sexes.(8) While it seems likely that dosage compensation is absent in Ceratitis capitata, if a system does exist, Tra apparently does not control it, since the XX pseudomales that result from loss of Tra are able to survive to adulthood. In fact, unlike Drosophila pseudomales, the Ceratitis pseudomales are fertile, confirming that the Ceratitis Y chromosome does not carry additional genes required for male development beyond the M factor.(27) Conclusions The results in this paper support the idea that genetic hierarchies evolve from the ‘‘bottom up’’ as proposed by Wilkins.(28) The slave gene at the bottom of the regulatory hierarchy, dsx, shows both structural and functional conservation across many species. At higher levels in the hierarchy, the regulatory genes can change from one species to the next. Thus although tra has maintained its role in regulating dsx, Sxl does not control tra. Instead, tra appears to have taken on the BioEssays 25.1 3 What the papers say role of the master switch gene that not only initiates the choice of sexual identity, but also maintains that choice through autoregulation. One issue that has been left unresolved is what controls the initial expression of tra in Ceratitis. Since maternal Cctra mRNAs that are spliced in the female pattern have been detected in unfertilized eggs, it is possible that protein translated from these mRNAs might be able to activate the initial alternative splicing of the zygotic transcripts in females. In males, the M factor might act to inhibit the maternal tra, perhaps by preventing its translation or by blocking the activity of the protein. In the future, it will be interesting to test this model and determine the identity and function of the M factor. In addition, it will be important to determine if the M factor functions transiently early in development to control male identity or whether it is present and is able to function throughout the entire lifecycle. Similarly, if Tra could be transiently expressed in XY embryos at a time when the M factor is no longer functional, the autoregulatory model predicts that it would initiate the female pathway and sex transform XY animals. References 1. Pane A, Salvemini M, Bovi PD, Polito C, Saccone G. The transformer gene in Ceratitis capitata provides a genetic basis for selecting and remembering the sexual fate. Development 2002;129:3715–3725. 2. Bell LR, Maine EM, Schedl P, Cline TW. Sex-lethal, a Drosophila sex determination switch gene, exhibits sex-specific RNA splicing and sequence similarity to RNA binding proteins. Cell 1988;55:1037– 1046. 3. Bell LR, Horabin JI, Schedl P, Cline TW. Positive autoregulation of sexlethal by alternative splicing maintains the female determined state in Drosophila. Cell 1991;65:229–239. 4. Jacobs PA, Strong JA. A case of human intersexuality having a possible XXY sex-determining mechanism. Nature 1959;183:302–303. 5. Ford CE, Jones KW, Polani PE, De Almeida JC, Briggs JH. A sexchromosome anomaly in a case of gonadal dysgenesis (Turner’s syndrome). Lancet 1959;1:711–713. 6. Welshons WJ, Russell LB. The Y chromosome as the bearer of male determining factors in the mouse. Proc Natl Acad Sci USA 1959;45:560–566. 7. Cline TW, Meyer BJ. Vive la difference:males vs females in flies vs worms. Annu Rev Genet 1996;30:637–702. 8. Schutt C, Nothiger R. Structure, function and evolution of sex-determining systems in Dipteran insects. Development 2000;127:667–677. 9. Hoshijima K, Inoue K, Higuchi I, Sakamoto H, Shimura Y. Control of doublesex alternative splicing by transformer and transformer-2 in Drosophila. Science 1991;252:833–836. 10. Bopp D, Calhoun G, Horabin JI, Samuels M, Schedl P. Sex-specific control of Sex-lethal is a conserved mechanism for sex determination in the genus Drosophila. Development 1996;122:971–982. 4 BioEssays 25.1 11. Meise M, Hilfiker-Kleiner D, Dubendorfer A, Brunner C, Nothiger R, Bopp D. Sex-lethal, the master sex-determining gene in Drosophila, is not sex- specifically regulated in Musca domestica. Development 1998; 125:1487–1494. 12. Saccone G, Peluso I, Artiaco D, Giordano E, Bopp D, Polito LC. The Ceratitis capitata homologue of the Drosophila sex-determining gene Sex-lethal is structurally conserved, but not sex-specifically regulated. Development 1998;125:1495–1500. 13. Sievert V, Kuhn S, Paululat A, Traut W. Sequence conservation and expression of the sex-lethal homologue in the fly Megaselia scalaris. Genome 2000;43:382–390. 14. Willhoeft U, Franz G. Identification of the sex-determining region of the Ceratitis capitata Y chromosome by deletion mapping. Genetics 1996; 144:737–745. 15. Kuhn S, Sievert V, Traut W. The sex-determining gene doublesex in the fly Megaselia scalaris:conserved structure and sex-specific splicing. Genome 2000;43:1011–1020. 16. Raymond CS, Shamu CE, Shen MM, Seifert KJ, Hirsch B, Hodgkin J, Zarkower D. Evidence for evolutionary conservation of sex-determining genes. Nature 1998;391:691–695. 17. Yi W, Zarkower D. Similarity of DNA binding and transcriptional regulation by Caenorhabditis elegans MAB-3 and Drosophila melanogaster DSX suggests conservation of sex determining mechanisms. Development 1999;126:873–881. 18. Lopez AJ. Alternative splicing of pre-mRNA:developmental consequences and mechanisms of regulation. Annu Rev Genet 1998;32: 279– 305. 19. Ryner LC, Baker BS. Regulation of doublesex pre-mRNA processing occurs by 30 -splice site activation. Genes Dev 1991;5:2071– 2085. 20. Heinrichs V, Ryner LC, Baker BS. Regulation of sex-specific selection of fruitless 50 splice sites by transformer and transformer-2. Mol Cell Biol 1998;18:450–458. 21. Kanopka A, Muhlemann O, Akusjarvi G. Inhibition by SR proteins of splicing of a regulated adenovirus pre-mRNA. Nature 1996;381:535– 538. 22. Bashaw GJ, Baker BS. The msl-2 dosage compensation gene of Drosophila encodes a putative DNA-binding protein whose expression is sex specifically regulated by Sex-lethal. Development 1995;121:3245– 3258. 23. Kelley RL, Solovyeva I, Lyman LM, Richman R, Solovyev V, Kuroda MI. Expression of msl-2 causes assembly of dosage compensation regulators on the X chromosomes and female lethality in Drosophila. Cell 1995;81:867–877. 24. Zhou S, et al. Male-specific lethal 2, a dosage compensation gene of Drosophila, undergoes sex-specific regulation and encodes a protein with a RING finger and a metallothionein-like cysteine cluster. EMBO J 1995;14:2884–2895. 25. Bernstein M, Cline TW. Differential effects of Sex-lethal mutations on dosage compensation early in Drosophila development. Genetics 1994; 136:1051–1061. 26. Kelley RL, Kuroda MI. Equality for X chromosomes. Science 1995; 270:1607–1610. 27. Schmidt R, Hediger M, Roth S, Nothiger R, Dubendorfer A. The Ychromosomal and autosomal male-determining M factors of Musca domestica are equivalent. Genetics 1997;147:271–280. 28. Wilkins AS. Moving up the hierarchy: a hypothesis on the evolution of a gentic sex determination pathway. Bioessays 1995;17:71–77.