Distinct functions of SR proteins in recruitment of U1 small nuclear ribonucleoprotein to alternative 5' splice sites (original) (raw)

Abstract

Alternative splicing of precursor messenger RNAs (pre-mRNAs) is an important mechanism for the regulation of gene expression. The members of the SR protein family of pre-mRNA splicing factors have distinct functions in promoting alternative splice site usage. Here we show that SR proteins are required for the first step of spliceosome assembly, interaction of the U1 small nuclear ribonucleoprotein complex (U1 snRNP) with the 5' splice site of the pre-mRNA. Further, we find that individual SR proteins have distinct abilities to promote interaction of U1 snRNP with alternative 5' splice junctions. These results suggest that SR proteins direct 5' splice site selection by regulation of U1 snRNP assembly onto the pre-mRNA.

2642

Images in this article

Selected References

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

  1. Cech T. R. The generality of self-splicing RNA: relationship to nuclear mRNA splicing. Cell. 1986 Jan 31;44(2):207–210. doi: 10.1016/0092-8674(86)90751-8. [DOI] [PubMed] [Google Scholar]
  2. Crispino J. D., Blencowe B. J., Sharp P. A. Complementation by SR proteins of pre-mRNA splicing reactions depleted of U1 snRNP. Science. 1994 Sep 23;265(5180):1866–1869. doi: 10.1126/science.8091213. [DOI] [PubMed] [Google Scholar]
  3. Eperon I. C., Ireland D. C., Smith R. A., Mayeda A., Krainer A. R. Pathways for selection of 5' splice sites by U1 snRNPs and SF2/ASF. EMBO J. 1993 Sep;12(9):3607–3617. doi: 10.1002/j.1460-2075.1993.tb06034.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fu X. D., Maniatis T. Isolation of a complementary DNA that encodes the mammalian splicing factor SC35. Science. 1992 Apr 24;256(5056):535–538. doi: 10.1126/science.1373910. [DOI] [PubMed] [Google Scholar]
  5. Fu X. D., Maniatis T. The 35-kDa mammalian splicing factor SC35 mediates specific interactions between U1 and U2 small nuclear ribonucleoprotein particles at the 3' splice site. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1725–1729. doi: 10.1073/pnas.89.5.1725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fu X. D. Specific commitment of different pre-mRNAs to splicing by single SR proteins. Nature. 1993 Sep 2;365(6441):82–85. doi: 10.1038/365082a0. [DOI] [PubMed] [Google Scholar]
  7. Ge H., Manley J. L. A protein factor, ASF, controls cell-specific alternative splicing of SV40 early pre-mRNA in vitro. Cell. 1990 Jul 13;62(1):25–34. doi: 10.1016/0092-8674(90)90236-8. [DOI] [PubMed] [Google Scholar]
  8. Ge H., Zuo P., Manley J. L. Primary structure of the human splicing factor ASF reveals similarities with Drosophila regulators. Cell. 1991 Jul 26;66(2):373–382. doi: 10.1016/0092-8674(91)90626-a. [DOI] [PubMed] [Google Scholar]
  9. Johnston R. F., Pickett S. C., Barker D. L. Autoradiography using storage phosphor technology. Electrophoresis. 1990 May;11(5):355–360. doi: 10.1002/elps.1150110503. [DOI] [PubMed] [Google Scholar]
  10. Kohtz J. D., Jamison S. F., Will C. L., Zuo P., Lührmann R., Garcia-Blanco M. A., Manley J. L. Protein-protein interactions and 5'-splice-site recognition in mammalian mRNA precursors. Nature. 1994 Mar 10;368(6467):119–124. doi: 10.1038/368119a0. [DOI] [PubMed] [Google Scholar]
  11. Krainer A. R., Mayeda A., Kozak D., Binns G. Functional expression of cloned human splicing factor SF2: homology to RNA-binding proteins, U1 70K, and Drosophila splicing regulators. Cell. 1991 Jul 26;66(2):383–394. doi: 10.1016/0092-8674(91)90627-b. [DOI] [PubMed] [Google Scholar]
  12. 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]
  13. Mayeda A., Zahler A. M., Krainer A. R., Roth M. B. Two members of a conserved family of nuclear phosphoproteins are involved in pre-mRNA splicing. Proc Natl Acad Sci U S A. 1992 Feb 15;89(4):1301–1304. doi: 10.1073/pnas.89.4.1301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Moore M. J., Sharp P. A. Site-specific modification of pre-mRNA: the 2'-hydroxyl groups at the splice sites. Science. 1992 May 15;256(5059):992–997. doi: 10.1126/science.1589782. [DOI] [PubMed] [Google Scholar]
  15. Nelson K. K., Green M. R. Mechanism for cryptic splice site activation during pre-mRNA splicing. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6253–6257. doi: 10.1073/pnas.87.16.6253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ner S. S., Atkinson T. C., Smith M. A method for introducing random single point deletions in specific DNA target sequences using oligonucleotides. Nucleic Acids Res. 1989 Jun 12;17(11):4015–4023. doi: 10.1093/nar/17.11.4015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sharp P. A. On the origin of RNA splicing and introns. Cell. 1985 Sep;42(2):397–400. doi: 10.1016/0092-8674(85)90092-3. [DOI] [PubMed] [Google Scholar]
  18. Sharp P. A. Split genes and RNA splicing. Cell. 1994 Jun 17;77(6):805–815. doi: 10.1016/0092-8674(94)90130-9. [DOI] [PubMed] [Google Scholar]
  19. Sontheimer E. J., Steitz J. A. The U5 and U6 small nuclear RNAs as active site components of the spliceosome. Science. 1993 Dec 24;262(5142):1989–1996. doi: 10.1126/science.8266094. [DOI] [PubMed] [Google Scholar]
  20. Steitz T. A., Steitz J. A. A general two-metal-ion mechanism for catalytic RNA. Proc Natl Acad Sci U S A. 1993 Jul 15;90(14):6498–6502. doi: 10.1073/pnas.90.14.6498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Tarn W. Y., Steitz J. A. SR proteins can compensate for the loss of U1 snRNP functions in vitro. Genes Dev. 1994 Nov 15;8(22):2704–2717. doi: 10.1101/gad.8.22.2704. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Treisman R., Orkin S. H., Maniatis T. Specific transcription and RNA splicing defects in five cloned beta-thalassaemia genes. Nature. 1983 Apr 14;302(5909):591–596. doi: 10.1038/302591a0. [DOI] [PubMed] [Google Scholar]
  25. Wu J. Y., Maniatis T. Specific interactions between proteins implicated in splice site selection and regulated alternative splicing. Cell. 1993 Dec 17;75(6):1061–1070. doi: 10.1016/0092-8674(93)90316-i. [DOI] [PubMed] [Google Scholar]
  26. Wyatt J. R., Sontheimer E. J., Steitz J. A. Site-specific cross-linking of mammalian U5 snRNP to the 5' splice site before the first step of pre-mRNA splicing. Genes Dev. 1992 Dec;6(12B):2542–2553. doi: 10.1101/gad.6.12b.2542. [DOI] [PubMed] [Google Scholar]
  27. Zahler A. M., Lane W. S., Stolk J. A., Roth M. B. SR proteins: a conserved family of pre-mRNA splicing factors. Genes Dev. 1992 May;6(5):837–847. doi: 10.1101/gad.6.5.837. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. Zahler A. M., Neugebauer K. M., Stolk J. A., Roth M. B. Human SR proteins and isolation of a cDNA encoding SRp75. Mol Cell Biol. 1993 Jul;13(7):4023–4028. doi: 10.1128/mcb.13.7.4023. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Zamore P. D., Green M. R. Biochemical characterization of U2 snRNP auxiliary factor: an essential pre-mRNA splicing factor with a novel intranuclear distribution. EMBO J. 1991 Jan;10(1):207–214. doi: 10.1002/j.1460-2075.1991.tb07937.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Zamore P. D., Patton J. G., Green M. R. Cloning and domain structure of the mammalian splicing factor U2AF. Nature. 1992 Feb 13;355(6361):609–614. doi: 10.1038/355609a0. [DOI] [PubMed] [Google Scholar]
  32. Zuo P., Manley J. L. The human splicing factor ASF/SF2 can specifically recognize pre-mRNA 5' splice sites. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3363–3367. doi: 10.1073/pnas.91.8.3363. [DOI] [PMC free article] [PubMed] [Google Scholar]