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. 1992 Sep;12(9):3939–3947. doi: 10.1128/mcb.12.9.3939

PRP38 encodes a yeast protein required for pre-mRNA splicing and maintenance of stable U6 small nuclear RNA levels.

S Blanton 1, A Srinivasan 1, B C Rymond 1
PMCID: PMC360275  PMID: 1508195

Abstract

An essential pre-mRNA splicing factor, the product of the PRP38 gene, has been genetically identified in a screen of temperature-sensitive mutants of Saccharomyces cerevisiae. Shifting temperature-sensitive prp38 cultures from 23 to 37 degrees C prevents the first cleavage-ligation event in the excision of introns from mRNA precursors. In vitro splicing inactivation and complementation studies suggest that the PRP38-encoded factor functions, at least in part, after stable splicing complex formation. The PRP38 locus contains a 726-bp open reading frame coding for an acidic 28-kDa polypeptide (PRP38). While PRP38 lacks obvious structural similarity to previously defined splicing factors, heat inactivation of PRP38, PRP19, or any of the known U6 (or U4/U6) small nuclear ribonucleoprotein-associating proteins (i.e., PRP3, PRP4, PRP6, and PRP24) leads to a common, unexpected consequence: intracellular U6 small nuclear RNA (snRNA) levels decrease as splicing activity is lost. Curiously, U4 snRNA, normally extensively base paired with U6 snRNA, persists in the virtual absence of U6 snRNA.

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Selected References

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  1. Abovich N., Legrain P., Rosbash M. The yeast PRP6 gene encodes a U4/U6 small nuclear ribonucleoprotein particle (snRNP) protein, and the PRP9 gene encodes a protein required for U2 snRNP binding. Mol Cell Biol. 1990 Dec;10(12):6417–6425. doi: 10.1128/mcb.10.12.6417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ares M., Jr U2 RNA from yeast is unexpectedly large and contains homology to vertebrate U4, U5, and U6 small nuclear RNAs. Cell. 1986 Oct 10;47(1):49–59. doi: 10.1016/0092-8674(86)90365-x. [DOI] [PubMed] [Google Scholar]
  3. Banroques J., Abelson J. N. PRP4: a protein of the yeast U4/U6 small nuclear ribonucleoprotein particle. Mol Cell Biol. 1989 Sep;9(9):3710–3719. doi: 10.1128/mcb.9.9.3710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Behrens S. E., Lührmann R. Immunoaffinity purification of a [U4/U6.U5] tri-snRNP from human cells. Genes Dev. 1991 Aug;5(8):1439–1452. doi: 10.1101/gad.5.8.1439. [DOI] [PubMed] [Google Scholar]
  5. Bjørn S. P., Soltyk A., Beggs J. D., Friesen J. D. PRP4 (RNA4) from Saccharomyces cerevisiae: its gene product is associated with the U4/U6 small nuclear ribonucleoprotein particle. Mol Cell Biol. 1989 Sep;9(9):3698–3709. doi: 10.1128/mcb.9.9.3698. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Broach J. R., Strathern J. N., Hicks J. B. Transformation in yeast: development of a hybrid cloning vector and isolation of the CAN1 gene. Gene. 1979 Dec;8(1):121–133. doi: 10.1016/0378-1119(79)90012-x. [DOI] [PubMed] [Google Scholar]
  7. Brow D. A., Guthrie C. Spliceosomal RNA U6 is remarkably conserved from yeast to mammals. Nature. 1988 Jul 21;334(6179):213–218. doi: 10.1038/334213a0. [DOI] [PubMed] [Google Scholar]
  8. Chang T. H., Clark M. W., Lustig A. J., Cusick M. E., Abelson J. RNA11 protein is associated with the yeast spliceosome and is localized in the periphery of the cell nucleus. Mol Cell Biol. 1988 Jun;8(6):2379–2393. doi: 10.1128/mcb.8.6.2379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chen J. H., Lin R. J. The yeast PRP2 protein, a putative RNA-dependent ATPase, shares extensive sequence homology with two other pre-mRNA splicing factors. Nucleic Acids Res. 1990 Nov 11;18(21):6447–6447. doi: 10.1093/nar/18.21.6447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cheng S. C., Abelson J. Fractionation and characterization of a yeast mRNA splicing extract. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2387–2391. doi: 10.1073/pnas.83.8.2387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cheng S. C., Abelson J. Spliceosome assembly in yeast. Genes Dev. 1987 Nov;1(9):1014–1027. doi: 10.1101/gad.1.9.1014. [DOI] [PubMed] [Google Scholar]
  12. Company M., Arenas J., Abelson J. Requirement of the RNA helicase-like protein PRP22 for release of messenger RNA from spliceosomes. Nature. 1991 Feb 7;349(6309):487–493. doi: 10.1038/349487a0. [DOI] [PubMed] [Google Scholar]
  13. Dalbadie-McFarland G., Abelson J. PRP5: a helicase-like protein required for mRNA splicing in yeast. Proc Natl Acad Sci U S A. 1990 Jun;87(11):4236–4240. doi: 10.1073/pnas.87.11.4236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Earnshaw W. C. Anionic regions in nuclear proteins. J Cell Biol. 1987 Oct;105(4):1479–1482. doi: 10.1083/jcb.105.4.1479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fabrizio P., Abelson J. Two domains of yeast U6 small nuclear RNA required for both steps of nuclear precursor messenger RNA splicing. Science. 1990 Oct 19;250(4979):404–409. doi: 10.1126/science.2145630. [DOI] [PubMed] [Google Scholar]
  16. Gietz R. D., Sugino A. New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene. 1988 Dec 30;74(2):527–534. doi: 10.1016/0378-1119(88)90185-0. [DOI] [PubMed] [Google Scholar]
  17. Green M. R. Biochemical mechanisms of constitutive and regulated pre-mRNA splicing. Annu Rev Cell Biol. 1991;7:559–599. doi: 10.1146/annurev.cb.07.110191.003015. [DOI] [PubMed] [Google Scholar]
  18. Guthrie C. Messenger RNA splicing in yeast: clues to why the spliceosome is a ribonucleoprotein. Science. 1991 Jul 12;253(5016):157–163. doi: 10.1126/science.1853200. [DOI] [PubMed] [Google Scholar]
  19. Hamilton R., Watanabe C. K., de Boer H. A. Compilation and comparison of the sequence context around the AUG startcodons in Saccharomyces cerevisiae mRNAs. Nucleic Acids Res. 1987 Apr 24;15(8):3581–3593. doi: 10.1093/nar/15.8.3581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kang W., Matsushita Y., Isono K. Cloning and analysis of YMR26, the nuclear gene for a mitochondrial ribosomal protein in Saccharomyces cerevisiae. Mol Gen Genet. 1991 Mar;225(3):474–482. doi: 10.1007/BF00261690. [DOI] [PubMed] [Google Scholar]
  22. Kretzner L., Rymond B. C., Rosbash M. S. cerevisiae U1 RNA is large and has limited primary sequence homology to metazoan U1 snRNA. Cell. 1987 Aug 14;50(4):593–602. doi: 10.1016/0092-8674(87)90032-8. [DOI] [PubMed] [Google Scholar]
  23. Krämer A., Keller W. Purification of a protein required for the splicing of pre-mRNA and its separation from the lariat debranching enzyme. EMBO J. 1985 Dec 16;4(13A):3571–3581. doi: 10.1002/j.1460-2075.1985.tb04119.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  25. Last R. L., Maddock J. R., Woolford J. L., Jr Evidence for related functions of the RNA genes of Saccharomyces cerevisiae. Genetics. 1987 Dec;117(4):619–631. doi: 10.1093/genetics/117.4.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lee W. C., Xue Z. X., Mélèse T. The NSR1 gene encodes a protein that specifically binds nuclear localization sequences and has two RNA recognition motifs. J Cell Biol. 1991 Apr;113(1):1–12. doi: 10.1083/jcb.113.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lin R. J., Lustig A. J., Abelson J. Splicing of yeast nuclear pre-mRNA in vitro requires a functional 40S spliceosome and several extrinsic factors. Genes Dev. 1987 Mar;1(1):7–18. doi: 10.1101/gad.1.1.7. [DOI] [PubMed] [Google Scholar]
  28. Lin R. J., Newman A. J., Cheng S. C., Abelson J. Yeast mRNA splicing in vitro. J Biol Chem. 1985 Nov 25;260(27):14780–14792. [PubMed] [Google Scholar]
  29. Lossky M., Anderson G. J., Jackson S. P., Beggs J. Identification of a yeast snRNP protein and detection of snRNP-snRNP interactions. Cell. 1987 Dec 24;51(6):1019–1026. doi: 10.1016/0092-8674(87)90588-5. [DOI] [PubMed] [Google Scholar]
  30. Lustig A. J., Lin R. J., Abelson J. The yeast RNA gene products are essential for mRNA splicing in vitro. Cell. 1986 Dec 26;47(6):953–963. doi: 10.1016/0092-8674(86)90810-x. [DOI] [PubMed] [Google Scholar]
  31. Lührmann R., Kastner B., Bach M. Structure of spliceosomal snRNPs and their role in pre-mRNA splicing. Biochim Biophys Acta. 1990 Nov 30;1087(3):265–292. doi: 10.1016/0167-4781(90)90001-i. [DOI] [PubMed] [Google Scholar]
  32. Madhani H. D., Bordonné R., Guthrie C. Multiple roles for U6 snRNA in the splicing pathway. Genes Dev. 1990 Dec;4(12B):2264–2277. doi: 10.1101/gad.4.12b.2264. [DOI] [PubMed] [Google Scholar]
  33. Moenne A., Camier S., Anderson G., Margottin F., Beggs J., Sentenac A. The U6 gene of Saccharomyces cerevisiae is transcribed by RNA polymerase C (III) in vivo and in vitro. EMBO J. 1990 Jan;9(1):271–277. doi: 10.1002/j.1460-2075.1990.tb08105.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Patterson B., Guthrie C. An essential yeast snRNA with a U5-like domain is required for splicing in vivo. Cell. 1987 Jun 5;49(5):613–624. doi: 10.1016/0092-8674(87)90537-x. [DOI] [PubMed] [Google Scholar]
  35. Pikielny C. W., Rosbash M. mRNA splicing efficiency in yeast and the contribution of nonconserved sequences. Cell. 1985 May;41(1):119–126. doi: 10.1016/0092-8674(85)90066-2. [DOI] [PubMed] [Google Scholar]
  36. Pikielny C. W., Rymond B. C., Rosbash M. Electrophoresis of ribonucleoproteins reveals an ordered assembly pathway of yeast splicing complexes. 1986 Nov 27-Dec 3Nature. 324(6095):341–345. doi: 10.1038/324341a0. [DOI] [PubMed] [Google Scholar]
  37. Rosbash M., Séraphin B. Who's on first? The U1 snRNP-5' splice site interaction and splicing. Trends Biochem Sci. 1991 May;16(5):187–190. doi: 10.1016/0968-0004(91)90073-5. [DOI] [PubMed] [Google Scholar]
  38. Rose M. D., Novick P., Thomas J. H., Botstein D., Fink G. R. A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector. Gene. 1987;60(2-3):237–243. doi: 10.1016/0378-1119(87)90232-0. [DOI] [PubMed] [Google Scholar]
  39. Ruby S. W., Abelson J. Pre-mRNA splicing in yeast. Trends Genet. 1991 Mar;7(3):79–85. doi: 10.1016/0168-9525(91)90276-V. [DOI] [PubMed] [Google Scholar]
  40. Ruskin B., Pikielny C. W., Rosbash M., Green M. R. Alternative branch points are selected during splicing of a yeast pre-mRNA in mammalian and yeast extracts. Proc Natl Acad Sci U S A. 1986 Apr;83(7):2022–2026. doi: 10.1073/pnas.83.7.2022. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rymond B. C., Rosbash M. A chemical modification/interference study of yeast pre-mRNA spliceosome assembly and splicing. Genes Dev. 1988 Apr;2(4):428–439. doi: 10.1101/gad.2.4.428. [DOI] [PubMed] [Google Scholar]
  42. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Schwer B., Guthrie C. PRP16 is an RNA-dependent ATPase that interacts transiently with the spliceosome. Nature. 1991 Feb 7;349(6309):494–499. doi: 10.1038/349494a0. [DOI] [PubMed] [Google Scholar]
  44. Shannon K. W., Guthrie C. Suppressors of a U4 snRNA mutation define a novel U6 snRNP protein with RNA-binding motifs. Genes Dev. 1991 May;5(5):773–785. doi: 10.1101/gad.5.5.773. [DOI] [PubMed] [Google Scholar]
  45. Sharp P. M., Tuohy T. M., Mosurski K. R. Codon usage in yeast: cluster analysis clearly differentiates highly and lowly expressed genes. Nucleic Acids Res. 1986 Jul 11;14(13):5125–5143. doi: 10.1093/nar/14.13.5125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Siliciano P. G., Brow D. A., Roiha H., Guthrie C. An essential snRNA from S. cerevisiae has properties predicted for U4, including interaction with a U6-like snRNA. Cell. 1987 Aug 14;50(4):585–592. doi: 10.1016/0092-8674(87)90031-6. [DOI] [PubMed] [Google Scholar]
  47. Siliciano P. G., Jones M. H., Guthrie C. Saccharomyces cerevisiae has a U1-like small nuclear RNA with unexpected properties. Science. 1987 Sep 18;237(4821):1484–1487. doi: 10.1126/science.3306922. [DOI] [PubMed] [Google Scholar]
  48. Strauss E. J., Guthrie C. A cold-sensitive mRNA splicing mutant is a member of the RNA helicase gene family. Genes Dev. 1991 Apr;5(4):629–641. doi: 10.1101/gad.5.4.629. [DOI] [PubMed] [Google Scholar]
  49. Séraphin B., Mattaj I. W. New pieces in the U5 puzzle. Curr Biol. 1991 Jun;1(3):147–149. doi: 10.1016/0960-9822(91)90215-i. [DOI] [PubMed] [Google Scholar]
  50. Teem J. L., Rosbash M. Expression of a beta-galactosidase gene containing the ribosomal protein 51 intron is sensitive to the rna2 mutation of yeast. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4403–4407. doi: 10.1073/pnas.80.14.4403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Vijayraghavan U., Abelson J. PRP18, a protein required for the second reaction in pre-mRNA splicing. Mol Cell Biol. 1990 Jan;10(1):324–332. doi: 10.1128/mcb.10.1.324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Vijayraghavan U., Company M., Abelson J. Isolation and characterization of pre-mRNA splicing mutants of Saccharomyces cerevisiae. Genes Dev. 1989 Aug;3(8):1206–1216. doi: 10.1101/gad.3.8.1206. [DOI] [PubMed] [Google Scholar]
  53. Whittaker E., Lossky M., Beggs J. D. Affinity purification of spliceosomes reveals that the precursor RNA processing protein PRP8, a protein in the U5 small nuclear ribonucleoprotein particle, is a component of yeast spliceosomes. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2216–2219. doi: 10.1073/pnas.87.6.2216. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Woolford J. L., Jr Nuclear pre-mRNA splicing in yeast. Yeast. 1989 Nov-Dec;5(6):439–457. doi: 10.1002/yea.320050604. [DOI] [PubMed] [Google Scholar]
  55. Xu Y., Petersen-Bjørn S., Friesen J. D. The PRP4 (RNA4) protein of Saccharomyces cerevisiae is associated with the 5' portion of the U4 small nuclear RNA. Mol Cell Biol. 1990 Mar;10(3):1217–1225. doi: 10.1128/mcb.10.3.1217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Yean S. L., Lin R. J. U4 small nuclear RNA dissociates from a yeast spliceosome and does not participate in the subsequent splicing reaction. Mol Cell Biol. 1991 Nov;11(11):5571–5577. doi: 10.1128/mcb.11.11.5571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Zieve G. W., Sauterer R. A. Cell biology of the snRNP particles. Crit Rev Biochem Mol Biol. 1990;25(1):1–46. doi: 10.3109/10409239009090604. [DOI] [PubMed] [Google Scholar]

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