Abstract
Strand exchange protein 1 (Sep1) (also referred to as exoribonuclease I [Xrn1]) from Saccharomyces cerevisiae has been implicated in DNA recombination, RNA turnover, karyogamy, and G4 DNA pairing among other disparate cellular processes. Using a genetic approach to study the role of SEP1/XRN1 in mitotic yeast cells, we identified mutations in the genes superkiller 2 (SKI2) and superkiller 3 (SKI3) as synthetically lethal with an sep1 null mutation. The SKI genes are thought to comprise an intracellular antiviral system controlling the expression of killer toxin from double-stranded RNA virus found in many yeast strains. However, the lethality of sep1 ski2 and sep1 ski3 mutants was independent of the L-A and M viruses, suggesting that the SKI genes act in a general cellular process in addition to virus control. We propose that Sep1/Xrn1 and Ski2 both act to block translation on transcripts targeted for degradation. Using a temperature-sensitive allele of SEP1/XRN1, we show that double mutants display a synthetic cell cycle arrest in late G1 at Start.
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- Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
- Amberg D. C., Goldstein A. L., Cole C. N. Isolation and characterization of RAT1: an essential gene of Saccharomyces cerevisiae required for the efficient nucleocytoplasmic trafficking of mRNA. Genes Dev. 1992 Jul;6(7):1173–1189. doi: 10.1101/gad.6.7.1173. [DOI] [PubMed] [Google Scholar]
- Blanc A., Ribas J. C., Wickner R. B., Sonenberg N. His-154 is involved in the linkage of the Saccharomyces cerevisiae L-A double-stranded RNA virus Gag protein to the cap structure of mRNAs and is essential for M1 satellite virus expression. Mol Cell Biol. 1994 Apr;14(4):2664–2674. doi: 10.1128/mcb.14.4.2664. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bähler J., Hagens G., Holzinger G., Scherthan H., Heyer W. D. Saccharomyces cerevisiae cells lacking the homologous pairing protein p175SEP1 arrest at pachytene during meiotic prophase. Chromosoma. 1994 Apr;103(2):129–141. doi: 10.1007/BF00352322. [DOI] [PubMed] [Google Scholar]
- Cao L., Alani E., Kleckner N. A pathway for generation and processing of double-strand breaks during meiotic recombination in S. cerevisiae. Cell. 1990 Jun 15;61(6):1089–1101. doi: 10.1016/0092-8674(90)90072-m. [DOI] [PubMed] [Google Scholar]
- Chen J., Kanaar R., Cozzarelli N. R. The Sep1 strand exchange protein from Saccharomyces cerevisiae promotes a paranemic joint between homologous DNA molecules. Genes Dev. 1994 Jun 1;8(11):1356–1366. doi: 10.1101/gad.8.11.1356. [DOI] [PubMed] [Google Scholar]
- Chiannilkulchai N., Stalder R., Riva M., Carles C., Werner M., Sentenac A. RPC82 encodes the highly conserved, third-largest subunit of RNA polymerase C (III) from Saccharomyces cerevisiae. Mol Cell Biol. 1992 Oct;12(10):4433–4440. doi: 10.1128/mcb.12.10.4433. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Davis B. D., Chen L. L., Tai P. C. Misread protein creates membrane channels: an essential step in the bactericidal action of aminoglycosides. Proc Natl Acad Sci U S A. 1986 Aug;83(16):6164–6168. doi: 10.1073/pnas.83.16.6164. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Decker C. J., Parker R. A turnover pathway for both stable and unstable mRNAs in yeast: evidence for a requirement for deadenylation. Genes Dev. 1993 Aug;7(8):1632–1643. doi: 10.1101/gad.7.8.1632. [DOI] [PubMed] [Google Scholar]
- Decker C. J., Parker R. Mechanisms of mRNA degradation in eukaryotes. Trends Biochem Sci. 1994 Aug;19(8):336–340. doi: 10.1016/0968-0004(94)90073-6. [DOI] [PubMed] [Google Scholar]
- Di Segni G., McConaughy B. L., Shapiro R. A., Aldrich T. L., Hall B. D. TAP1, a yeast gene that activates the expression of a tRNA gene with a defective internal promoter. Mol Cell Biol. 1993 Jun;13(6):3424–3433. doi: 10.1128/mcb.13.6.3424. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dihanich M., van Tuinen E., Lambris J. D., Marshallsay B. Accumulation of viruslike particles in a yeast mutant lacking a mitochondrial pore protein. Mol Cell Biol. 1989 Mar;9(3):1100–1108. doi: 10.1128/mcb.9.3.1100. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Donovan W. P., Kushner S. R. Polynucleotide phosphorylase and ribonuclease II are required for cell viability and mRNA turnover in Escherichia coli K-12. Proc Natl Acad Sci U S A. 1986 Jan;83(1):120–124. doi: 10.1073/pnas.83.1.120. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Drebot M. A., Johnston G. C., Friesen J. D., Singer R. A. An impaired RNA polymerase II activity in Saccharomyces cerevisiae causes cell-cycle inhibition at START. Mol Gen Genet. 1993 Nov;241(3-4):327–334. doi: 10.1007/BF00284685. [DOI] [PubMed] [Google Scholar]
- Dykstra C. C., Hamatake R. K., Sugino A. DNA strand transfer protein beta from yeast mitotic cells differs from strand transfer protein alpha from meiotic cells. J Biol Chem. 1990 Jul 5;265(19):10968–10973. [PubMed] [Google Scholar]
- Dykstra C. C., Kitada K., Clark A. B., Hamatake R. K., Sugino A. Cloning and characterization of DST2, the gene for DNA strand transfer protein beta from Saccharomyces cerevisiae. Mol Cell Biol. 1991 May;11(5):2583–2592. doi: 10.1128/mcb.11.5.2583. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fried H. M., Fink G. R. Electron microscopic heteroduplex analysis of "killer" double-stranded RNA species from yeast. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4224–4228. doi: 10.1073/pnas.75.9.4224. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gietz D., St Jean A., Woods R. A., Schiestl R. H. Improved method for high efficiency transformation of intact yeast cells. Nucleic Acids Res. 1992 Mar 25;20(6):1425–1425. doi: 10.1093/nar/20.6.1425. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guarente L. Synthetic enhancement in gene interaction: a genetic tool come of age. Trends Genet. 1993 Oct;9(10):362–366. doi: 10.1016/0168-9525(93)90042-g. [DOI] [PubMed] [Google Scholar]
- Henry Y., Wood H., Morrissey J. P., Petfalski E., Kearsey S., Tollervey D. The 5' end of yeast 5.8S rRNA is generated by exonucleases from an upstream cleavage site. EMBO J. 1994 May 15;13(10):2452–2463. doi: 10.1002/j.1460-2075.1994.tb06530.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Heyer W. D., Evans D. H., Kolodner R. D. Renaturation of DNA by a Saccharomyces cerevisiae protein that catalyzes homologous pairing and strand exchange. J Biol Chem. 1988 Oct 15;263(29):15189–15195. [PubMed] [Google Scholar]
- Heyer W. D., Johnson A. W., Reinhart U., Kolodner R. D. Regulation and intracellular localization of Saccharomyces cerevisiae strand exchange protein 1 (Sep1/Xrn1/Kem1), a multifunctional exonuclease. Mol Cell Biol. 1995 May;15(5):2728–2736. doi: 10.1128/mcb.15.5.2728. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hsu C. L., Stevens A. Yeast cells lacking 5'-->3' exoribonuclease 1 contain mRNA species that are poly(A) deficient and partially lack the 5' cap structure. Mol Cell Biol. 1993 Aug;13(8):4826–4835. doi: 10.1128/mcb.13.8.4826. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Iizuka N., Najita L., Franzusoff A., Sarnow P. Cap-dependent and cap-independent translation by internal initiation of mRNAs in cell extracts prepared from Saccharomyces cerevisiae. Mol Cell Biol. 1994 Nov;14(11):7322–7330. doi: 10.1128/mcb.14.11.7322. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Johnson A. W., Kolodner R. D. Strand exchange protein 1 from Saccharomyces cerevisiae. A novel multifunctional protein that contains DNA strand exchange and exonuclease activities. J Biol Chem. 1991 Jul 25;266(21):14046–14054. [PubMed] [Google Scholar]
- Kearsey S., Kipling D. Recombination and RNA processing: a common strand? Trends Cell Biol. 1991 Nov;1(5):110–112. doi: 10.1016/0962-8924(91)90101-e. [DOI] [PubMed] [Google Scholar]
- Kenna M., Stevens A., McCammon M., Douglas M. G. An essential yeast gene with homology to the exonuclease-encoding XRN1/KEM1 gene also encodes a protein with exoribonuclease activity. Mol Cell Biol. 1993 Jan;13(1):341–350. doi: 10.1128/mcb.13.1.341. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kim J., Ljungdahl P. O., Fink G. R. kem mutations affect nuclear fusion in Saccharomyces cerevisiae. Genetics. 1990 Dec;126(4):799–812. doi: 10.1093/genetics/126.4.799. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kolodner R., Evans D. H., Morrison P. T. Purification and characterization of an activity from Saccharomyces cerevisiae that catalyzes homologous pairing and strand exchange. Proc Natl Acad Sci U S A. 1987 Aug;84(16):5560–5564. doi: 10.1073/pnas.84.16.5560. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kowalczykowski S. C., Eggleston A. K. Homologous pairing and DNA strand-exchange proteins. Annu Rev Biochem. 1994;63:991–1043. doi: 10.1146/annurev.bi.63.070194.005015. [DOI] [PubMed] [Google Scholar]
- Kranz J. E., Holm C. Cloning by function: an alternative approach for identifying yeast homologs of genes from other organisms. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6629–6633. doi: 10.1073/pnas.87.17.6629. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Larimer F. W., Hsu C. L., Maupin M. K., Stevens A. Characterization of the XRN1 gene encoding a 5'-->3' exoribonuclease: sequence data and analysis of disparate protein and mRNA levels of gene-disrupted yeast cells. Gene. 1992 Oct 12;120(1):51–57. doi: 10.1016/0378-1119(92)90008-d. [DOI] [PubMed] [Google Scholar]
- Liu Z., Gilbert W. The yeast KEM1 gene encodes a nuclease specific for G4 tetraplex DNA: implication of in vivo functions for this novel DNA structure. Cell. 1994 Jul 1;77(7):1083–1092. doi: 10.1016/0092-8674(94)90447-2. [DOI] [PubMed] [Google Scholar]
- Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
- Matsumoto Y., Fishel R., Wickner R. B. Circular single-stranded RNA replicon in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7628–7632. doi: 10.1073/pnas.87.19.7628. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Muhlrad D., Decker C. J., Parker R. Deadenylation of the unstable mRNA encoded by the yeast MFA2 gene leads to decapping followed by 5'-->3' digestion of the transcript. Genes Dev. 1994 Apr 1;8(7):855–866. doi: 10.1101/gad.8.7.855. [DOI] [PubMed] [Google Scholar]
- Muhlrad D., Parker R. Premature translational termination triggers mRNA decapping. Nature. 1994 Aug 18;370(6490):578–581. doi: 10.1038/370578a0. [DOI] [PubMed] [Google Scholar]
- Munroe D., Jacobson A. Tales of poly(A): a review. Gene. 1990 Jul 16;91(2):151–158. doi: 10.1016/0378-1119(90)90082-3. [DOI] [PubMed] [Google Scholar]
- Nasmyth K. Control of the yeast cell cycle by the Cdc28 protein kinase. Curr Opin Cell Biol. 1993 Apr;5(2):166–179. doi: 10.1016/0955-0674(93)90099-c. [DOI] [PubMed] [Google Scholar]
- Nemeroff M. E., Bruenn J. A. Initiation by the yeast viral transcriptase in vitro. J Biol Chem. 1987 May 15;262(14):6785–6787. [PubMed] [Google Scholar]
- Proweller A., Butler S. Efficient translation of poly(A)-deficient mRNAs in Saccharomyces cerevisiae. Genes Dev. 1994 Nov 1;8(21):2629–2640. doi: 10.1101/gad.8.21.2629. [DOI] [PubMed] [Google Scholar]
- Reed S. I. The selection of S. cerevisiae mutants defective in the start event of cell division. Genetics. 1980 Jul;95(3):561–577. doi: 10.1093/genetics/95.3.561. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Reenan R. A., Kolodner R. D. Characterization of insertion mutations in the Saccharomyces cerevisiae MSH1 and MSH2 genes: evidence for separate mitochondrial and nuclear functions. Genetics. 1992 Dec;132(4):975–985. doi: 10.1093/genetics/132.4.975. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rhee S. K., Icho T., Wickner R. B. Structure and nuclear localization signal of the SKI3 antiviral protein of Saccharomyces cerevisiae. Yeast. 1989 May-Jun;5(3):149–158. doi: 10.1002/yea.320050304. [DOI] [PubMed] [Google Scholar]
- Ridley S. P., Sommer S. S., Wickner R. B. Superkiller mutations in Saccharomyces cerevisiae suppress exclusion of M2 double-stranded RNA by L-A-HN and confer cold sensitivity in the presence of M and L-A-HN. Mol Cell Biol. 1984 Apr;4(4):761–770. doi: 10.1128/mcb.4.4.761. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sachs A. B., Davis R. W. Translation initiation and ribosomal biogenesis: involvement of a putative rRNA helicase and RPL46. Science. 1990 Mar 2;247(4946):1077–1079. doi: 10.1126/science.2408148. [DOI] [PubMed] [Google Scholar]
- Stevens A. An exoribonuclease from Saccharomyces cerevisiae: effect of modifications of 5' end groups on the hydrolysis of substrates to 5' mononucleotides. Biochem Biophys Res Commun. 1978 Mar 30;81(2):656–661. doi: 10.1016/0006-291x(78)91586-3. [DOI] [PubMed] [Google Scholar]
- Stevens A., Hsu C. L., Isham K. R., Larimer F. W. Fragments of the internal transcribed spacer 1 of pre-rRNA accumulate in Saccharomyces cerevisiae lacking 5'----3' exoribonuclease 1. J Bacteriol. 1991 Nov;173(21):7024–7028. doi: 10.1128/jb.173.21.7024-7028.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stevens A. Purification and characterization of a Saccharomyces cerevisiae exoribonuclease which yields 5'-mononucleotides by a 5' leads to 3' mode of hydrolysis. J Biol Chem. 1980 Apr 10;255(7):3080–3085. [PubMed] [Google Scholar]
- Symington L. S., Fogarty L. M., Kolodner R. Genetic recombination of homologous plasmids catalyzed by cell-free extracts of Saccharomyces cerevisiae. Cell. 1983 Dec;35(3 Pt 2):805–813. doi: 10.1016/0092-8674(83)90113-7. [DOI] [PubMed] [Google Scholar]
- Tishkoff D. X., Johnson A. W., Kolodner R. D. Molecular and genetic analysis of the gene encoding the Saccharomyces cerevisiae strand exchange protein Sep1. Mol Cell Biol. 1991 May;11(5):2593–2608. doi: 10.1128/mcb.11.5.2593. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tishkoff D. X., Rockmill B., Roeder G. S., Kolodner R. D. The sep1 mutant of Saccharomyces cerevisiae arrests in pachytene and is deficient in meiotic recombination. Genetics. 1995 Feb;139(2):495–509. doi: 10.1093/genetics/139.2.495. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Toh-E A., Guerry P., Wickner R. B. Chromosomal superkiller mutants of Saccharomyces cerevisiae. J Bacteriol. 1978 Dec;136(3):1002–1007. doi: 10.1128/jb.136.3.1002-1007.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wickner R. B. Double-stranded and single-stranded RNA viruses of Saccharomyces cerevisiae. Annu Rev Microbiol. 1992;46:347–375. doi: 10.1146/annurev.mi.46.100192.002023. [DOI] [PubMed] [Google Scholar]
- Wickner R. B. Host function of MAK16: G1 arrest by a mak16 mutant of Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1988 Aug;85(16):6007–6011. doi: 10.1073/pnas.85.16.6007. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wickner R. B., Leibowitz M. J. Two chromosomal genes required for killing expression in killer strains of Saccharomyces cerevisiae. Genetics. 1976 Mar 25;82(3):429–442. doi: 10.1093/genetics/82.3.429. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Widner W. R., Wickner R. B. Evidence that the SKI antiviral system of Saccharomyces cerevisiae acts by blocking expression of viral mRNA. Mol Cell Biol. 1993 Jul;13(7):4331–4341. doi: 10.1128/mcb.13.7.4331. [DOI] [PMC free article] [PubMed] [Google Scholar]