Skip to main content
RNA logoLink to RNA
. 2000 Jan;6(1):111–120. doi: 10.1017/s1355838200991982

The SRm160/300 splicing coactivator subunits.

B J Blencowe 1, G Baurén 1, A G Eldridge 1, R Issner 1, J A Nickerson 1, E Rosonina 1, P A Sharp 1
PMCID: PMC1369899  PMID: 10668804

Abstract

The SRm160/300 splicing coactivator, which consists of the serine/arginine (SR)-related nuclear matrix protein of 160 kDa and a 300-kDa nuclear matrix antigen, functions in splicing by promoting critical interactions between splicing factors bound to pre-mRNA, including snRNPs and SR family proteins. In this article we report the isolation of a cDNA encoding the 300-kDa antigen and investigate the activity of it and SRm160 in splicing. Like SRm160, the 300-kDa antigen contains domains rich in alternating S and R residues but lacks an RNA recognition motif; the protein is accordingly named "SRm300." SRm300 also contains a novel and highly conserved N-terminal domain, several unique repeated motifs rich in S, R, and proline residues, and two very long polyserine tracts. Surprisingly, specific depletion of SRm300 does not prevent the splicing of pre-mRNAs shown previously to require SRm160/300. Addition of recombinant SRm160 alone to SRm160/300-depleted reactions specifically activates splicing. The results indicate that SRm160 may be the more critical component of the SRm160/300 coactivator in the splicing of SRm160/300-dependent pre-mRNAs.

Full Text

The Full Text of this article is available as a PDF (725.2 KB).

Selected References

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

  1. Beil B., Screaton G., Stamm S. Molecular cloning of htra2-beta-1 and htra2-beta-2, two human homologs of tra-2 generated by alternative splicing. DNA Cell Biol. 1997 Jun;16(6):679–690. doi: 10.1089/dna.1997.16.679. [DOI] [PubMed] [Google Scholar]
  2. Blencowe B. J., Bowman J. A., McCracken S., Rosonina E. SR-related proteins and the processing of messenger RNA precursors. Biochem Cell Biol. 1999;77(4):277–291. [PubMed] [Google Scholar]
  3. Blencowe B. J., Issner R., Kim J., Mccaw P., Sharp P. A. New proteins related to the Ser-Arg family of splicing factors. RNA. 1995 Oct;1(8):852–865. [PMC free article] [PubMed] [Google Scholar]
  4. Blencowe B. J., Issner R., Nickerson J. A., Sharp P. A. A coactivator of pre-mRNA splicing. Genes Dev. 1998 Apr 1;12(7):996–1009. doi: 10.1101/gad.12.7.996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Blencowe B. J., Nickerson J. A., Issner R., Penman S., Sharp P. A. Association of nuclear matrix antigens with exon-containing splicing complexes. J Cell Biol. 1994 Nov;127(3):593–607. doi: 10.1083/jcb.127.3.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Blencowe B. J., Ouzounis C. A. The PWI motif: a new protein domain in splicing factors. Trends Biochem Sci. 1999 May;24(5):179–180. doi: 10.1016/s0968-0004(99)01387-0. [DOI] [PubMed] [Google Scholar]
  7. Bourquin J. P., Stagljar I., Meier P., Moosmann P., Silke J., Baechi T., Georgiev O., Schaffner W. A serine/arginine-rich nuclear matrix cyclophilin interacts with the C-terminal domain of RNA polymerase II. Nucleic Acids Res. 1997 Jun 1;25(11):2055–2061. doi: 10.1093/nar/25.11.2055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brendel V., Bucher P., Nourbakhsh I. R., Blaisdell B. E., Karlin S. Methods and algorithms for statistical analysis of protein sequences. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2002–2006. doi: 10.1073/pnas.89.6.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Colwill K., Pawson T., Andrews B., Prasad J., Manley J. L., Bell J. C., Duncan P. I. The Clk/Sty protein kinase phosphorylates SR splicing factors and regulates their intranuclear distribution. EMBO J. 1996 Jan 15;15(2):265–275. [PMC free article] [PubMed] [Google Scholar]
  10. Cáceres J. F., Stamm S., Helfman D. M., Krainer A. R. Regulation of alternative splicing in vivo by overexpression of antagonistic splicing factors. Science. 1994 Sep 16;265(5179):1706–1709. doi: 10.1126/science.8085156. [DOI] [PubMed] [Google Scholar]
  11. Dauwalder B., Amaya-Manzanares F., Mattox W. A human homologue of the Drosophila sex determination factor transformer-2 has conserved splicing regulatory functions. Proc Natl Acad Sci U S A. 1996 Aug 20;93(17):9004–9009. doi: 10.1073/pnas.93.17.9004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Denhez F., Lafyatis R. Conservation of regulated alternative splicing and identification of functional domains in vertebrate homologs to the Drosophila splicing regulator, suppressor-of-white-apricot. J Biol Chem. 1994 Jun 10;269(23):16170–16179. [PubMed] [Google Scholar]
  13. Dignam J. D., Lebovitz R. M., Roeder R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. doi: 10.1093/nar/11.5.1475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dirksen W. P., Hampson R. K., Sun Q., Rottman F. M. A purine-rich exon sequence enhances alternative splicing of bovine growth hormone pre-mRNA. J Biol Chem. 1994 Mar 4;269(9):6431–6436. [PubMed] [Google Scholar]
  15. Eldridge A. G., Li Y., Sharp P. A., Blencowe B. J. The SRm160/300 splicing coactivator is required for exon-enhancer function. Proc Natl Acad Sci U S A. 1999 May 25;96(11):6125–6130. doi: 10.1073/pnas.96.11.6125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fu X. D., Mayeda A., Maniatis T., Krainer A. R. General splicing factors SF2 and SC35 have equivalent activities in vitro, and both affect alternative 5' and 3' splice site selection. Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11224–11228. doi: 10.1073/pnas.89.23.11224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fu X. D. The superfamily of arginine/serine-rich splicing factors. RNA. 1995 Sep;1(7):663–680. [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. Gui J. F., Lane W. S., Fu X. D. A serine kinase regulates intracellular localization of splicing factors in the cell cycle. Nature. 1994 Jun 23;369(6482):678–682. doi: 10.1038/369678a0. [DOI] [PubMed] [Google Scholar]
  20. Gui J. F., Tronchère H., Chandler S. D., Fu X. D. Purification and characterization of a kinase specific for the serine- and arginine-rich pre-mRNA splicing factors. Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):10824–10828. doi: 10.1073/pnas.91.23.10824. [DOI] [PMC free article] [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. Krainer A. R., Conway G. C., Kozak D. Purification and characterization of pre-mRNA splicing factor SF2 from HeLa cells. Genes Dev. 1990 Jul;4(7):1158–1171. doi: 10.1101/gad.4.7.1158. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. 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]
  25. Lopez A. J. Alternative splicing of pre-mRNA: developmental consequences and mechanisms of regulation. Annu Rev Genet. 1998;32:279–305. doi: 10.1146/annurev.genet.32.1.279. [DOI] [PubMed] [Google Scholar]
  26. Manley J. L., Tacke R. SR proteins and splicing control. Genes Dev. 1996 Jul 1;10(13):1569–1579. doi: 10.1101/gad.10.13.1569. [DOI] [PubMed] [Google Scholar]
  27. Mayeda A., Badolato J., Kobayashi R., Zhang M. Q., Gardiner E. M., Krainer A. R. Purification and characterization of human RNPS1: a general activator of pre-mRNA splicing. EMBO J. 1999 Aug 16;18(16):4560–4570. doi: 10.1093/emboj/18.16.4560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Meier U. T., Blobel G. Nopp140 shuttles on tracks between nucleolus and cytoplasm. Cell. 1992 Jul 10;70(1):127–138. doi: 10.1016/0092-8674(92)90539-o. [DOI] [PubMed] [Google Scholar]
  29. Misteli T. RNA splicing: What has phosphorylation got to do with it? Curr Biol. 1999 Mar 25;9(6):R198–R200. doi: 10.1016/s0960-9822(99)80128-6. [DOI] [PubMed] [Google Scholar]
  30. Nagase T., Ishikawa K., Nakajima D., Ohira M., Seki N., Miyajima N., Tanaka A., Kotani H., Nomura N., Ohara O. Prediction of the coding sequences of unidentified human genes. VII. The complete sequences of 100 new cDNA clones from brain which can code for large proteins in vitro. DNA Res. 1997 Apr 28;4(2):141–150. doi: 10.1093/dnares/4.2.141. [DOI] [PubMed] [Google Scholar]
  31. Roscigno R. F., Garcia-Blanco M. A. SR proteins escort the U4/U6.U5 tri-snRNP to the spliceosome. RNA. 1995 Sep;1(7):692–706. [PMC free article] [PubMed] [Google Scholar]
  32. Spikes D. A., Kramer J., Bingham P. M., Van Doren K. SWAP pre-mRNA splicing regulators are a novel, ancient protein family sharing a highly conserved sequence motif with the prp21 family of constitutive splicing proteins. Nucleic Acids Res. 1994 Oct 25;22(21):4510–4519. doi: 10.1093/nar/22.21.4510. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. 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]
  34. Tacke R., Tohyama M., Ogawa S., Manley J. L. Human Tra2 proteins are sequence-specific activators of pre-mRNA splicing. Cell. 1998 Apr 3;93(1):139–148. doi: 10.1016/s0092-8674(00)81153-8. [DOI] [PubMed] [Google Scholar]
  35. Tanaka K., Watakabe A., Shimura Y. Polypurine sequences within a downstream exon function as a splicing enhancer. Mol Cell Biol. 1994 Feb;14(2):1347–1354. doi: 10.1128/mcb.14.2.1347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. 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]
  37. Tian M., Maniatis T. A splicing enhancer exhibits both constitutive and regulated activities. Genes Dev. 1994 Jul 15;8(14):1703–1712. doi: 10.1101/gad.8.14.1703. [DOI] [PubMed] [Google Scholar]
  38. Valcárcel J., Green M. R. The SR protein family: pleiotropic functions in pre-mRNA splicing. Trends Biochem Sci. 1996 Aug;21(8):296–301. [PubMed] [Google Scholar]
  39. Wan K. M., Nickerson J. A., Krockmalnic G., Penman S. The B1C8 protein is in the dense assemblies of the nuclear matrix and relocates to the spindle and pericentriolar filaments at mitosis. Proc Natl Acad Sci U S A. 1994 Jan 18;91(2):594–598. doi: 10.1073/pnas.91.2.594. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wang J., Manley J. L. Overexpression of the SR proteins ASF/SF2 and SC35 influences alternative splicing in vivo in diverse ways. RNA. 1995 May;1(3):335–346. [PMC free article] [PubMed] [Google Scholar]
  41. Yeakley J. M., Morfin J. P., Rosenfeld M. G., Fu X. D. A complex of nuclear proteins mediates SR protein binding to a purine-rich splicing enhancer. Proc Natl Acad Sci U S A. 1996 Jul 23;93(15):7582–7587. doi: 10.1073/pnas.93.15.7582. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Yuryev A., Patturajan M., Litingtung Y., Joshi R. V., Gentile C., Gebara M., Corden J. L. The C-terminal domain of the largest subunit of RNA polymerase II interacts with a novel set of serine/arginine-rich proteins. Proc Natl Acad Sci U S A. 1996 Jul 9;93(14):6975–6980. doi: 10.1073/pnas.93.14.6975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. 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]
  44. Zhang W. J., Wu J. Y. Functional properties of p54, a novel SR protein active in constitutive and alternative splicing. Mol Cell Biol. 1996 Oct;16(10):5400–5408. doi: 10.1128/mcb.16.10.5400. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Zhang W. J., Wu J. Y. Sip1, a novel RS domain-containing protein essential for pre-mRNA splicing. Mol Cell Biol. 1998 Feb;18(2):676–684. doi: 10.1128/mcb.18.2.676. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. de la Cruz J., Kressler D., Linder P. Unwinding RNA in Saccharomyces cerevisiae: DEAD-box proteins and related families. Trends Biochem Sci. 1999 May;24(5):192–198. doi: 10.1016/s0968-0004(99)01376-6. [DOI] [PubMed] [Google Scholar]

Articles from RNA are provided here courtesy of The RNA Society

RESOURCES