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. 1996 Aug 20;93(17):9004–9009. doi: 10.1073/pnas.93.17.9004

A human homologue of the Drosophila sex determination factor transformer-2 has conserved splicing regulatory functions.

B Dauwalder 1, F Amaya-Manzanares 1, W Mattox 1
PMCID: PMC38585  PMID: 8799144

Abstract

Regulation of gene expression through alternative pre-mRNA splicing appears to occur in all metazoans, but most of our knowledge about splicing regulators derives from studies on genetically identified factors from Drosophila. Among the best studied of these is the transformer-2 (TRA-2) protein which, in combination with the transformer (TRA) protein, directs sex-specific splicing of pre-mRNA from the sex determination gene doublesex (dsx). Here we report the identification of htra-2 alpha, a human homologue of tra-2. Two alternative types of htra-2 alpha cDNA clones were identified that encode different protein isoforms with striking organizational similarity to Drosophila tra-2 proteins. When expressed in flies, one hTRA-2 alpha isoform partially replaces the function of Drosophila TRA-2, affecting both female sexual differentiation and alternative splicing of dsx pre-mRNA. Like Drosophila TRA-2, the ability of hTRA-2 alpha to regulate dsx is female-specific and depends on the presence of the dsx splicing enhancer. These results demonstrate that htra-2 alpha has conserved a striking degree of functional specificity during evolution and leads us to suggest that, although they are likely to serve different roles in development, the tra-2 products of flies and humans have similar molecular functions.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amrein H., Gorman M., Nöthiger R. The sex-determining gene tra-2 of Drosophila encodes a putative RNA binding protein. Cell. 1988 Dec 23;55(6):1025–1035. doi: 10.1016/0092-8674(88)90247-4. [DOI] [PubMed] [Google Scholar]
  2. Amrein H., Hedley M. L., Maniatis T. The role of specific protein-RNA and protein-protein interactions in positive and negative control of pre-mRNA splicing by Transformer 2. Cell. 1994 Feb 25;76(4):735–746. doi: 10.1016/0092-8674(94)90512-6. [DOI] [PubMed] [Google Scholar]
  3. Amrein H., Maniatis T., Nöthiger R. Alternatively spliced transcripts of the sex-determining gene tra-2 of Drosophila encode functional proteins of different size. EMBO J. 1990 Nov;9(11):3619–3629. doi: 10.1002/j.1460-2075.1990.tb07573.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Baker B. S., Ridge K. A. Sex and the single cell. I. On the action of major loci affecting sex determination in Drosophila melanogaster. Genetics. 1980 Feb;94(2):383–423. doi: 10.1093/genetics/94.2.383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Baker B. S., Wolfner M. F. A molecular analysis of doublesex, a bifunctional gene that controls both male and female sexual differentiation in Drosophila melanogaster. Genes Dev. 1988 Apr;2(4):477–489. doi: 10.1101/gad.2.4.477. [DOI] [PubMed] [Google Scholar]
  6. Bell L. R., Maine E. M., Schedl P., Cline T. W. Sex-lethal, a Drosophila sex determination switch gene, exhibits sex-specific RNA splicing and sequence similarity to RNA binding proteins. Cell. 1988 Dec 23;55(6):1037–1046. doi: 10.1016/0092-8674(88)90248-6. [DOI] [PubMed] [Google Scholar]
  7. Belote J. M., Baker B. S. The dual functions of a sex determination gene in Drosophila melanogaster. Dev Biol. 1983 Feb;95(2):512–517. doi: 10.1016/0012-1606(83)90054-4. [DOI] [PubMed] [Google Scholar]
  8. Birney E., Kumar S., Krainer A. R. Analysis of the RNA-recognition motif and RS and RGG domains: conservation in metazoan pre-mRNA splicing factors. Nucleic Acids Res. 1993 Dec 25;21(25):5803–5816. doi: 10.1093/nar/21.25.5803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Burtis K. C., Baker B. S. Drosophila doublesex gene controls somatic sexual differentiation by producing alternatively spliced mRNAs encoding related sex-specific polypeptides. Cell. 1989 Mar 24;56(6):997–1010. doi: 10.1016/0092-8674(89)90633-8. [DOI] [PubMed] [Google Scholar]
  10. Cline T. W. The affairs of daughterless and the promiscuity of developmental regulators. Cell. 1989 Oct 20;59(2):231–234. doi: 10.1016/0092-8674(89)90280-8. [DOI] [PubMed] [Google Scholar]
  11. Cronmiller C., Schedl P., Cline T. W. Molecular characterization of daughterless, a Drosophila sex determination gene with multiple roles in development. Genes Dev. 1988 Dec;2(12A):1666–1676. doi: 10.1101/gad.2.12a.1666. [DOI] [PubMed] [Google Scholar]
  12. Flickinger T. W., Salz H. K. The Drosophila sex determination gene snf encodes a nuclear protein with sequence and functional similarity to the mammalian U1A snRNP protein. Genes Dev. 1994 Apr 15;8(8):914–925. doi: 10.1101/gad.8.8.914. [DOI] [PubMed] [Google Scholar]
  13. Fu X. D. The superfamily of arginine/serine-rich splicing factors. RNA. 1995 Sep;1(7):663–680. [PMC free article] [PubMed] [Google Scholar]
  14. Goralski T. J., Edström J. E., Baker B. S. The sex determination locus transformer-2 of Drosophila encodes a polypeptide with similarity to RNA binding proteins. Cell. 1989 Mar 24;56(6):1011–1018. doi: 10.1016/0092-8674(89)90634-x. [DOI] [PubMed] [Google Scholar]
  15. Hazelrigg T., Tu C. Sex-specific processing of the Drosophila exuperantia transcript is regulated in male germ cells by the tra-2 gene. Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10752–10756. doi: 10.1073/pnas.91.22.10752. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hedley M. L., Maniatis T. Sex-specific splicing and polyadenylation of dsx pre-mRNA requires a sequence that binds specifically to tra-2 protein in vitro. Cell. 1991 May 17;65(4):579–586. doi: 10.1016/0092-8674(91)90090-l. [DOI] [PubMed] [Google Scholar]
  17. Heinrichs V., Baker B. S. The Drosophila SR protein RBP1 contributes to the regulation of doublesex alternative splicing by recognizing RBP1 RNA target sequences. EMBO J. 1995 Aug 15;14(16):3987–4000. doi: 10.1002/j.1460-2075.1995.tb00070.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hodgkin J. Genetic sex determination mechanisms and evolution. Bioessays. 1992 Apr;14(4):253–261. doi: 10.1002/bies.950140409. [DOI] [PubMed] [Google Scholar]
  19. Hoshijima K., Inoue K., Higuchi I., Sakamoto H., Shimura Y. Control of doublesex alternative splicing by transformer and transformer-2 in Drosophila. Science. 1991 May 10;252(5007):833–836. doi: 10.1126/science.1902987. [DOI] [PubMed] [Google Scholar]
  20. Ingham P. W. Signalling by hedgehog family proteins in Drosophila and vertebrate development. Curr Opin Genet Dev. 1995 Aug;5(4):492–498. doi: 10.1016/0959-437x(95)90054-k. [DOI] [PubMed] [Google Scholar]
  21. Inoue K., Hoshijima K., Higuchi I., Sakamoto H., Shimura Y. Binding of the Drosophila transformer and transformer-2 proteins to the regulatory elements of doublesex primary transcript for sex-specific RNA processing. Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):8092–8096. doi: 10.1073/pnas.89.17.8092. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Johnson R. L., Tabin C. The long and short of hedgehog signaling. Cell. 1995 May 5;81(3):313–316. doi: 10.1016/0092-8674(95)90381-x. [DOI] [PubMed] [Google Scholar]
  23. Kenan D. J., Query C. C., Keene J. D. RNA recognition: towards identifying determinants of specificity. Trends Biochem Sci. 1991 Jun;16(6):214–220. doi: 10.1016/0968-0004(91)90088-d. [DOI] [PubMed] [Google Scholar]
  24. Krainer A. R., Conway G. C., Kozak D. The essential pre-mRNA splicing factor SF2 influences 5' splice site selection by activating proximal sites. Cell. 1990 Jul 13;62(1):35–42. doi: 10.1016/0092-8674(90)90237-9. [DOI] [PubMed] [Google Scholar]
  25. Lavigueur A., La Branche H., Kornblihtt A. R., Chabot B. A splicing enhancer in the human fibronectin alternate ED1 exon interacts with SR proteins and stimulates U2 snRNP binding. Genes Dev. 1993 Dec;7(12A):2405–2417. doi: 10.1101/gad.7.12a.2405. [DOI] [PubMed] [Google Scholar]
  26. Lennon G., Auffray C., Polymeropoulos M., Soares M. B. The I.M.A.G.E. Consortium: an integrated molecular analysis of genomes and their expression. Genomics. 1996 Apr 1;33(1):151–152. doi: 10.1006/geno.1996.0177. [DOI] [PubMed] [Google Scholar]
  27. Lin C. H., Patton J. G. Regulation of alternative 3' splice site selection by constitutive splicing factors. RNA. 1995 May;1(3):234–245. [PMC free article] [PubMed] [Google Scholar]
  28. Lynch K. W., Maniatis T. Synergistic interactions between two distinct elements of a regulated splicing enhancer. Genes Dev. 1995 Feb 1;9(3):284–293. doi: 10.1101/gad.9.3.284. [DOI] [PubMed] [Google Scholar]
  29. Mattox W., Baker B. S. Autoregulation of the splicing of transcripts from the transformer-2 gene of Drosophila. Genes Dev. 1991 May;5(5):786–796. doi: 10.1101/gad.5.5.786. [DOI] [PubMed] [Google Scholar]
  30. Mattox W., McGuffin M. E., Baker B. S. A negative feedback mechanism revealed by functional analysis of the alternative isoforms of the Drosophila splicing regulator transformer-2. Genetics. 1996 May;143(1):303–314. doi: 10.1093/genetics/143.1.303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mattox W., Palmer M. J., Baker B. S. Alternative splicing of the sex determination gene transformer-2 is sex-specific in the germ line but not in the soma. Genes Dev. 1990 May;4(5):789–805. doi: 10.1101/gad.4.5.789. [DOI] [PubMed] [Google Scholar]
  32. McKeown M., Belote J. M., Baker B. S. A molecular analysis of transformer, a gene in Drosophila melanogaster that controls female sexual differentiation. Cell. 1987 Feb 13;48(3):489–499. doi: 10.1016/0092-8674(87)90199-1. [DOI] [PubMed] [Google Scholar]
  33. McKeown M., Belote J. M., Boggs R. T. Ectopic expression of the female transformer gene product leads to female differentiation of chromosomally male Drosophila. Cell. 1988 Jun 17;53(6):887–895. doi: 10.1016/s0092-8674(88)90369-8. [DOI] [PubMed] [Google Scholar]
  34. Murre C., McCaw P. S., Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989 Mar 10;56(5):777–783. doi: 10.1016/0092-8674(89)90682-x. [DOI] [PubMed] [Google Scholar]
  35. Nagoshi R. N., Baker B. S. Regulation of sex-specific RNA splicing at the Drosophila doublesex gene: cis-acting mutations in exon sequences alter sex-specific RNA splicing patterns. Genes Dev. 1990 Jan;4(1):89–97. doi: 10.1101/gad.4.1.89. [DOI] [PubMed] [Google Scholar]
  36. Nagoshi R. N., McKeown M., Burtis K. C., Belote J. M., Baker B. S. The control of alternative splicing at genes regulating sexual differentiation in D. melanogaster. Cell. 1988 Apr 22;53(2):229–236. doi: 10.1016/0092-8674(88)90384-4. [DOI] [PubMed] [Google Scholar]
  37. Neiman A. M. Conservation and reiteration of a kinase cascade. Trends Genet. 1993 Nov;9(11):390–394. doi: 10.1016/0168-9525(93)90139-9. [DOI] [PubMed] [Google Scholar]
  38. Peng X., Mount S. M. Genetic enhancement of RNA-processing defects by a dominant mutation in B52, the Drosophila gene for an SR protein splicing factor. Mol Cell Biol. 1995 Nov;15(11):6273–6282. doi: 10.1128/mcb.15.11.6273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Peterson C., Legerski R. High-frequency transformation of human repair-deficient cell lines by an Epstein-Barr virus-based cDNA expression vector. Gene. 1991 Nov 15;107(2):279–284. doi: 10.1016/0378-1119(91)90328-9. [DOI] [PubMed] [Google Scholar]
  40. Polycarpou-Schwarz M., Gunderson S. I., Kandels-Lewis S., Seraphin B., Mattaj I. W. Drosophila SNF/D25 combines the functions of the two snRNP proteins U1A and U2B' that are encoded separately in human, potato, and yeast. RNA. 1996 Jan;2(1):11–23. [PMC free article] [PubMed] [Google Scholar]
  41. Ramchatesingh J., Zahler A. M., Neugebauer K. M., Roth M. B., Cooper T. A. A subset of SR proteins activates splicing of the cardiac troponin T alternative exon by direct interactions with an exonic enhancer. Mol Cell Biol. 1995 Sep;15(9):4898–4907. doi: 10.1128/mcb.15.9.4898. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Ryner L. C., Baker B. S. Regulation of doublesex pre-mRNA processing occurs by 3'-splice site activation. Genes Dev. 1991 Nov;5(11):2071–2085. doi: 10.1101/gad.5.11.2071. [DOI] [PubMed] [Google Scholar]
  43. Ryner L. C., Swain A. Sex in the '90s. Cell. 1995 May 19;81(4):483–493. doi: 10.1016/0092-8674(95)90069-1. [DOI] [PubMed] [Google Scholar]
  44. Scherly D., Boelens W., Dathan N. A., van Venrooij W. J., Mattaj I. W. Major determinants of the specificity of interaction between small nuclear ribonucleoproteins U1A and U2B'' and their cognate RNAs. Nature. 1990 Jun 7;345(6275):502–506. doi: 10.1038/345502a0. [DOI] [PubMed] [Google Scholar]
  45. Sun Q., Mayeda A., Hampson R. K., Krainer A. R., Rottman F. M. General splicing factor SF2/ASF promotes alternative splicing by binding to an exonic splicing enhancer. Genes Dev. 1993 Dec;7(12B):2598–2608. doi: 10.1101/gad.7.12b.2598. [DOI] [PubMed] [Google Scholar]
  46. Tian M., Maniatis T. A splicing enhancer complex controls alternative splicing of doublesex pre-mRNA. Cell. 1993 Jul 16;74(1):105–114. doi: 10.1016/0092-8674(93)90298-5. [DOI] [PubMed] [Google Scholar]
  47. Tian M., Maniatis T. Positive control of pre-mRNA splicing in vitro. Science. 1992 Apr 10;256(5054):237–240. doi: 10.1126/science.1566072. [DOI] [PubMed] [Google Scholar]
  48. Zachar Z., Chou T. B., Bingham P. M. Evidence that a regulatory gene autoregulates splicing of its transcript. EMBO J. 1987 Dec 20;6(13):4105–4111. doi: 10.1002/j.1460-2075.1987.tb02756.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Zahler A. M., Neugebauer K. M., Lane W. S., Roth M. B. Distinct functions of SR proteins in alternative pre-mRNA splicing. Science. 1993 Apr 9;260(5105):219–222. doi: 10.1126/science.8385799. [DOI] [PubMed] [Google Scholar]

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