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. 2004 Feb;166(2):729–739. doi: 10.1534/genetics.166.2.729

Identification of Edc3p as an enhancer of mRNA decapping in Saccharomyces cerevisiae.

Meenakshi Kshirsagar 1, Roy Parker 1
PMCID: PMC1470743  PMID: 15020463

Abstract

The major pathway of mRNA decay in yeast initiates with deadenylation, followed by mRNA decapping and 5'-3' exonuclease digestion. An in silico approach was used to identify new proteins involved in the mRNA decay pathway. One such protein, Edc3p, was identified as a conserved protein of unknown function having extensive two-hybrid interactions with several proteins involved in mRNA decapping and 5'-3' degradation including Dcp1p, Dcp2p, Dhh1p, Lsm1p, and the 5'-3' exonuclease, Xrn1p. We show that Edc3p can stimulate mRNA decapping of both unstable and stable mRNAs in yeast when the decapping enzyme is compromised by temperature-sensitive alleles of either the DCP1 or the DCP2 genes. In these cases, deletion of EDC3 caused a synergistic mRNA-decapping defect at the permissive temperatures. The edc3Delta had no effect when combined with the lsm1Delta, dhh1Delta, or pat1Delta mutations, which appear to affect an early step in the decapping pathway. This suggests that Edc3p specifically affects the function of the decapping enzyme per se. Consistent with a functional role in decapping, GFP-tagged Edc3p localizes to cytoplasmic foci involved in mRNA decapping referred to as P-bodies. These results identify Edc3p as a new protein involved in the decapping reaction.

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Selected References

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  1. 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]
  2. Anderson J. S., Parker R. P. The 3' to 5' degradation of yeast mRNAs is a general mechanism for mRNA turnover that requires the SKI2 DEVH box protein and 3' to 5' exonucleases of the exosome complex. EMBO J. 1998 Mar 2;17(5):1497–1506. doi: 10.1093/emboj/17.5.1497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Beelman C. A., Stevens A., Caponigro G., LaGrandeur T. E., Hatfield L., Fortner D. M., Parker R. An essential component of the decapping enzyme required for normal rates of mRNA turnover. Nature. 1996 Aug 15;382(6592):642–646. doi: 10.1038/382642a0. [DOI] [PubMed] [Google Scholar]
  4. Bonnerot C., Boeck R., Lapeyre B. The two proteins Pat1p (Mrt1p) and Spb8p interact in vivo, are required for mRNA decay, and are functionally linked to Pab1p. Mol Cell Biol. 2000 Aug;20(16):5939–5946. doi: 10.1128/mcb.20.16.5939-5946.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bouveret E., Rigaut G., Shevchenko A., Wilm M., Séraphin B. A Sm-like protein complex that participates in mRNA degradation. EMBO J. 2000 Apr 3;19(7):1661–1671. doi: 10.1093/emboj/19.7.1661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Caponigro G., Muhlrad D., Parker R. A small segment of the MAT alpha 1 transcript promotes mRNA decay in Saccharomyces cerevisiae: a stimulatory role for rare codons. Mol Cell Biol. 1993 Sep;13(9):5141–5148. doi: 10.1128/mcb.13.9.5141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Coller J. M., Tucker M., Sheth U., Valencia-Sanchez M. A., Parker R. The DEAD box helicase, Dhh1p, functions in mRNA decapping and interacts with both the decapping and deadenylase complexes. RNA. 2001 Dec;7(12):1717–1727. doi: 10.1017/s135583820101994x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Daugeron M. C., Mauxion F., Séraphin B. The yeast POP2 gene encodes a nuclease involved in mRNA deadenylation. Nucleic Acids Res. 2001 Jun 15;29(12):2448–2455. doi: 10.1093/nar/29.12.2448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fischer Nicole, Weis Karsten. The DEAD box protein Dhh1 stimulates the decapping enzyme Dcp1. EMBO J. 2002 Jun 3;21(11):2788–2797. doi: 10.1093/emboj/21.11.2788. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fromont-Racine M., Mayes A. E., Brunet-Simon A., Rain J. C., Colley A., Dix I., Decourty L., Joly N., Ricard F., Beggs J. D. Genome-wide protein interaction screens reveal functional networks involving Sm-like proteins. Yeast. 2000 Jun 30;17(2):95–110. doi: 10.1002/1097-0061(20000630)17:2<95::AID-YEA16>3.0.CO;2-H. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fromont-Racine M., Mayes A. E., Brunet-Simon A., Rain J. C., Colley A., Dix I., Decourty L., Joly N., Ricard F., Beggs J. D. Genome-wide protein interaction screens reveal functional networks involving Sm-like proteins. Yeast. 2000 Jun 30;17(2):95–110. doi: 10.1002/1097-0061(20000630)17:2<95::AID-YEA16>3.0.CO;2-H. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fromont-Racine M., Rain J. C., Legrain P. Toward a functional analysis of the yeast genome through exhaustive two-hybrid screens. Nat Genet. 1997 Jul;16(3):277–282. doi: 10.1038/ng0797-277. [DOI] [PubMed] [Google Scholar]
  13. Gavin Anne-Claude, Bösche Markus, Krause Roland, Grandi Paola, Marzioch Martina, Bauer Andreas, Schultz Jörg, Rick Jens M., Michon Anne-Marie, Cruciat Cristina-Maria. Functional organization of the yeast proteome by systematic analysis of protein complexes. Nature. 2002 Jan 10;415(6868):141–147. doi: 10.1038/415141a. [DOI] [PubMed] [Google Scholar]
  14. Giaever Guri, Chu Angela M., Ni Li, Connelly Carla, Riles Linda, Véronneau Steeve, Dow Sally, Lucau-Danila Ankuta, Anderson Keith, André Bruno. Functional profiling of the Saccharomyces cerevisiae genome. Nature. 2002 Jul 25;418(6896):387–391. doi: 10.1038/nature00935. [DOI] [PubMed] [Google Scholar]
  15. Hatfield L., Beelman C. A., Stevens A., Parker R. Mutations in trans-acting factors affecting mRNA decapping in Saccharomyces cerevisiae. Mol Cell Biol. 1996 Oct;16(10):5830–5838. doi: 10.1128/mcb.16.10.5830. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. He F., Peltz S. W., Donahue J. L., Rosbash M., Jacobson A. Stabilization and ribosome association of unspliced pre-mRNAs in a yeast upf1- mutant. Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):7034–7038. doi: 10.1073/pnas.90.15.7034. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. He W., Parker R. The yeast cytoplasmic LsmI/Pat1p complex protects mRNA 3' termini from partial degradation. Genetics. 2001 Aug;158(4):1445–1455. doi: 10.1093/genetics/158.4.1445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hilleren P., Parker R. Mechanisms of mRNA surveillance in eukaryotes. Annu Rev Genet. 1999;33:229–260. doi: 10.1146/annurev.genet.33.1.229. [DOI] [PubMed] [Google Scholar]
  19. Hishigaki H., Nakai K., Ono T., Tanigami A., Takagi T. Assessment of prediction accuracy of protein function from protein--protein interaction data. Yeast. 2001 Apr;18(6):523–531. doi: 10.1002/yea.706. [DOI] [PubMed] [Google Scholar]
  20. Ingelfinger Dierk, Arndt-Jovin Donna J., Lührmann Reinhard, Achsel Tilmann. The human LSm1-7 proteins colocalize with the mRNA-degrading enzymes Dcp1/2 and Xrnl in distinct cytoplasmic foci. RNA. 2002 Dec;8(12):1489–1501. [PMC free article] [PubMed] [Google Scholar]
  21. Longtine M. S., McKenzie A., 3rd, Demarini D. J., Shah N. G., Wach A., Brachat A., Philippsen P., Pringle J. R. Additional modules for versatile and economical PCR-based gene deletion and modification in Saccharomyces cerevisiae. Yeast. 1998 Jul;14(10):953–961. doi: 10.1002/(SICI)1097-0061(199807)14:10<953::AID-YEA293>3.0.CO;2-U. [DOI] [PubMed] [Google Scholar]
  22. Lykke-Andersen Jens. Identification of a human decapping complex associated with hUpf proteins in nonsense-mediated decay. Mol Cell Biol. 2002 Dec;22(23):8114–8121. doi: 10.1128/MCB.22.23.8114-8121.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Muhlrad D., Parker R. Mutations affecting stability and deadenylation of the yeast MFA2 transcript. Genes Dev. 1992 Nov;6(11):2100–2111. doi: 10.1101/gad.6.11.2100. [DOI] [PubMed] [Google Scholar]
  25. Schwartz D. C., Parker R. mRNA decapping in yeast requires dissociation of the cap binding protein, eukaryotic translation initiation factor 4E. Mol Cell Biol. 2000 Nov;20(21):7933–7942. doi: 10.1128/mcb.20.21.7933-7942.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schwartz David, Decker Carolyn J., Parker Roy. The enhancer of decapping proteins, Edc1p and Edc2p, bind RNA and stimulate the activity of the decapping enzyme. RNA. 2003 Feb;9(2):239–251. doi: 10.1261/rna.2171203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Schwikowski B., Uetz P., Fields S. A network of protein-protein interactions in yeast. Nat Biotechnol. 2000 Dec;18(12):1257–1261. doi: 10.1038/82360. [DOI] [PubMed] [Google Scholar]
  28. Sheth Ujwal, Parker Roy. Decapping and decay of messenger RNA occur in cytoplasmic processing bodies. Science. 2003 May 2;300(5620):805–808. doi: 10.1126/science.1082320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Steiger Michelle A., Parker Roy. Analyzing mRNA decay in Saccharomyces cerevisiae. Methods Enzymol. 2002;351:648–660. doi: 10.1016/s0076-6879(02)51875-9. [DOI] [PubMed] [Google Scholar]
  30. Steiger Michelle, Carr-Schmid Anne, Schwartz David C., Kiledjian Megerditch, Parker Roy. Analysis of recombinant yeast decapping enzyme. RNA. 2003 Feb;9(2):231–238. doi: 10.1261/rna.2151403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Steiger Michelle, Carr-Schmid Anne, Schwartz David C., Kiledjian Megerditch, Parker Roy. Analysis of recombinant yeast decapping enzyme. RNA. 2003 Feb;9(2):231–238. doi: 10.1261/rna.2151403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Tharun S., He W., Mayes A. E., Lennertz P., Beggs J. D., Parker R. Yeast Sm-like proteins function in mRNA decapping and decay. Nature. 2000 Mar 30;404(6777):515–518. doi: 10.1038/35006676. [DOI] [PubMed] [Google Scholar]
  33. Tharun S., Parker R. Analysis of mutations in the yeast mRNA decapping enzyme. Genetics. 1999 Apr;151(4):1273–1285. doi: 10.1093/genetics/151.4.1273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tharun S., Parker R. Targeting an mRNA for decapping: displacement of translation factors and association of the Lsm1p-7p complex on deadenylated yeast mRNAs. Mol Cell. 2001 Nov;8(5):1075–1083. doi: 10.1016/s1097-2765(01)00395-1. [DOI] [PubMed] [Google Scholar]
  35. Thompson J. D., Higgins D. G., Gibson T. J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994 Nov 11;22(22):4673–4680. doi: 10.1093/nar/22.22.4673. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Tucker M., Valencia-Sanchez M. A., Staples R. R., Chen J., Denis C. L., Parker R. The transcription factor associated Ccr4 and Caf1 proteins are components of the major cytoplasmic mRNA deadenylase in Saccharomyces cerevisiae. Cell. 2001 Feb 9;104(3):377–386. doi: 10.1016/s0092-8674(01)00225-2. [DOI] [PubMed] [Google Scholar]
  37. Tucker Morgan, Staples Robin R., Valencia-Sanchez Marco A., Muhlrad Denise, Parker Roy. Ccr4p is the catalytic subunit of a Ccr4p/Pop2p/Notp mRNA deadenylase complex in Saccharomyces cerevisiae. EMBO J. 2002 Mar 15;21(6):1427–1436. doi: 10.1093/emboj/21.6.1427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Uetz P., Giot L., Cagney G., Mansfield T. A., Judson R. S., Knight J. R., Lockshon D., Narayan V., Srinivasan M., Pochart P. A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisiae. Nature. 2000 Feb 10;403(6770):623–627. doi: 10.1038/35001009. [DOI] [PubMed] [Google Scholar]
  39. Wilusz C. J., Wang W., Peltz S. W. Curbing the nonsense: the activation and regulation of mRNA surveillance. Genes Dev. 2001 Nov 1;15(21):2781–2785. doi: 10.1101/gad.943701. [DOI] [PubMed] [Google Scholar]
  40. Zuk D., Belk J. P., Jacobson A. Temperature-sensitive mutations in the Saccharomyces cerevisiae MRT4, GRC5, SLA2 and THS1 genes result in defects in mRNA turnover. Genetics. 1999 Sep;153(1):35–47. doi: 10.1093/genetics/153.1.35. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. van Dijk Erwin, Cougot Nicolas, Meyer Sylke, Babajko Sylvie, Wahle Elmar, Séraphin Bertrand. Human Dcp2: a catalytically active mRNA decapping enzyme located in specific cytoplasmic structures. EMBO J. 2002 Dec 16;21(24):6915–6924. doi: 10.1093/emboj/cdf678. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. van Hoof A., Staples R. R., Baker R. E., Parker R. Function of the ski4p (Csl4p) and Ski7p proteins in 3'-to-5' degradation of mRNA. Mol Cell Biol. 2000 Nov;20(21):8230–8243. doi: 10.1128/mcb.20.21.8230-8243.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]

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