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2000 | 47 | 1 | 181-190
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Suppressors of translation initiation defect in hem12 locus of Saccharomyces cerevisiae.

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A system for the positive selection of transational initiation suppressors in S. cerevisiae has been developed. A mutant with an ATA initiation codon in the HEM12 gene, encoding uroporphyrinogen decarboxylase, was used to select cis- and trans-acting suppressors. These suppressors partially restore growth on nonfermentable carbon sources, such as glycerol, but still allow the accumulation of porphyrins. All extragenic suppressors are mapped to the SUI1 locus, encoding initiation factor eIF1. The effect of the hem12 mutation is also partially reversed by the known SUI3 suppressor encoding the β subunit of eIF2. In contrast, the sui2 suppressor encoding the α subunit of eIF2 does not affect the hem 12 phenotype. The intragenic suppressors are able to restore the translation of hem12 due to the generation of additional, in frame AUG codons upstream of the hem12-14 mutation. Mutational analysis of the HEM12 leader sequence was also performed to determine the role of small open reading frames (uORFs) present upstream of the HEM12 ORF. Studies on the expression of integrated hem12-1/4-lacZ fusion, devoid of all upstream ATGs, indicate a lack of regulatory effect of uORFs on HEM12 translation.
Physical description
  • Department of Genetics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, A. Pawińskiego 5a, 02-106 Warszawa, Poland
  • Department of Genetics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, A. Pawińskiego 5a, 02-106 Warszawa, Poland
  • Department of Genetics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, A. Pawińskiego 5a, 02-106 Warszawa, Poland
  • Department of Genetics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, A. Pawińskiego 5a, 02-106 Warszawa, Poland
  • 1. Merric, W.C. & Hershey, J.W.B. (1996) The pathway and mechanism of eucaryotic protein synthesis: Translational control (Hershey, J.W.B., Matthews, M.B. & Sonnenberg, N., eds.) pp. 31-69, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
  • 2. Kozak, M. (1999) Initiation of translation in procaryotes and eucaryotes. Gene 234, 187-208.
  • 3. Donahue, T.F. & Cigan, A.M. (1988) Genetic selection for mutations that reduce or abolish ribosomal recognition of the HIS4 translational initiator region. Mol. Cell. Biol. 8, 2955-2963.
  • 4. Castilho-Valavicius, B., Yoon, H. & Donahue, T.F. (1990) Genetic characterization of the Saccharomyces cerevisiae translational initiation suppressors sui1, sui2 and SUI3 and their effects on HIS4 expression. Genetics 124, 483-495.
  • 5. Yoon, H.J. & Donahue, T.F. (1992) The sui1 suppressor locus in Saccharomyces cerevisiae encodes the translation factor that functions during Met-tRNAi recognition of the start codon. Mol. Cell. Biol. 12, 248-260.
  • 6. Kasperaitis, M.A.M., Vooma, H.O. & Thomas, A.A.M. (1995) The amino acid sequence of eucaryotic translation initiation factor 1 and its similarity to yeast initiation factor SUI1. FEBS Lett. 365, 47-50.
  • 7. Narada, T., MacMillan, S.E., Donahue, T.F., Polekhina, G., Reshetnikova, L., Clark, B.F.C. & Nyborg, J. (1996) SUI1/p16 is required for the activity of eucaryotic translation initiation factor 3 in Saccharomyces cerevisiae. Mol. Cell. Biol. 16, 2307-2313.
  • 8. Cui, Y., Dinman, J.D., Kinzy, T.G. & Peltz, S.W. (1998) The Mof2/Sui1 protein is a general monitor of translational accuracy. Mol. Cell. Biol. 18, 1506-1516.
  • 9. Cigan, A.M., Pabich, E.K., Feng, L. & Donahue, T.F. (1989) Yeast translation initiation suppressor sui2 encodes the α subunit of eucaryotic factor 2 and shares identity with the human α subunit. Proc. Natl. Acad. Sci U.S.A. 86, 2784-2788.
  • 10. Donahue, T.F., Cigan, A.M., Pabich, E.K. & Castilho-Valavicius, B. (1988) Mutations at a Zn(II) finger motif in the yeast eIF-2β gene alter ribosomal start-site selection during the scanning process. Cell 54, 621-632.
  • 11. Dorris, D.R., Ericson, F.L. & Hannig, E.M. (1995) Mutations in GCD11, the structural gene for eIF-2γ in yeast, alter translation regulation of GCN4 and the selection of the start site for protein synthesis. EMBO J. 14, 2239-2249.
  • 12. Cigan, A.M., Feng, L. & Donahue, T.F. (1988) tRNAMet functions in directing the scanning ribosome to the start site of translation. Science 242, 93-97.
  • 13. McCarthy, J.E.G. (1998) Posttranslational control of gene expression in yeast. Microbiol. Mol. Biol. Rev. 62, 1492-1553.
  • 14. Hinnebusch, A.G. (1997) Traslational regulation of yeast GCN4. J. Biol. Chem. 272, 21661-21664.
  • 15. Garey, J.R., Labbe-Bois, R., Chelstowska, A., Rytka, J., Harrison, L., Kushner, J. & Labbe, P. (1992) Uroporphyrin decarboxylase in Saccharomyces cerevisiae: HEM12 gene sequence and evidence for two conserved glycines essential for enzymatic activity. Eur. J. Biochem. 205, 1011-1016.
  • 16. Labbe-Bois, R. & Labbe, P. (1990) Tetrapyrrole and heme biosynthesis in the yeast Saccharomyces cerevisiae; in Biosynthesis of Heme and Chlorophylls (Dailey, H.A., ed.) pp. 235-285, McGraw-Hill Book Co., New York.
  • 17. Chełstowska, A., Żołądek, T., Garey, J., Kushner, J., Rytka, J. & Labbe-Bois, R. (1992) Identification of amino acid changes affecting yeast uroporphyrinogen decarboxylase activity by sequence analysis of hem12 mutant alleles. Biochem. J. 288, 753-757.
  • 18. Kurlandzka, A. & Rytka, J. (1985) Mutants of Saccharomyces cerevisiae partially defective in the last steps of the haem biosynthetic pathway: Isolation and genetical characterization. J. Gen. Microbiol. 131, 2909-2918.
  • 19. Żołądek, T., Chełstowska, A., Labbe-Bois, R. & Rytka, J. (1995) Isolation and characterization of the extragenic mutations affecting the expression of the uroporphyrinogen decarboxylase gene (HEM12) in Saccharomyces cerevisiae. Mol. Gen. Genet. 247, 471-481.
  • 20. Chen, D.C., Yang, B.C. & Kuo, T.T. (1992) One-step transformation of yeast in stationary phase. Curr. Genet. 21, 83-84.
  • 21. Rose, M.D., Winston, F. & Hieter, P. (1990) Methods in Yeast Genetics: a Laboratory Course Manual. Cold Spring Harbor Laboratory Press, New York.
  • 22. Chien, C.T., Bartel, P.L., Sternglanz, R. & Fields, S. (1991) The two-hybrid system: A method to identify and clone genes for proteins that interact with a protein of interest. Proc. Natl. Acad. Sci. U.S.A. 88, 9578-9582.
  • 23. Parent, S.A., Fenimore, C.M. & Bostian, K.A. (1985) Vector systems for the expression, analysis and cloning of DNA sequences in S. cerevisiae. Yeast 1, 83-138.
  • 24. Myers, A.M., Tzagoloff, A., Kinney, D.M. & Lusty, C.J. (1986) Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions. Gene 45, 299-310.
  • 25. Sambrook, J., Fritsch, E.F. & Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, New York.
  • 26. Rytka, J., Biliński, T. & Labbe-Bois, R. (1984) Modified uroporphyrinogen decarboxylase activity in a yeast mutant which mimics porphyria cutanea tarda. Biochem. J. 218, 405-413.
  • 27. Volland, C. & Urban-Grimal, D. (1988) The presequence of yeast 5-aminolevulinate synthase is not required for targeting to mitochondria. J. Biol. Chem. 263, 8294-8299.
  • 28. Felix, F. & Brouillet, N. (1990) Purification and properties of uroporphyrinogen decarboxylase from Saccharomyces cerevisiae. Eur. J. Biochem. 188, 393-403.
  • 29. Cui, Y., González, C.I., Kinzy, T.G., Dinman, J. D. & Peltz, S.W. (1999) Mutations in the MOF2/SUI1 gene affect both translation and nonsense-mediated mRNA decay. RNA 5, 794-804.
  • 30. Aravind, L. & Koonin, E.V. (1999) Novel predicted RNA-binding domains associated with the translation machinery. J. Mol. Evol. 48, 291-302.
  • 31. Dever, T.E., Feng, L., Wek, R.C., Cigan, A.M., Donahue, T.F. & Hinnebusch, A.G. (1992) Phosphorylation of initiation factor 2α by protein kinase GCN2 mediates gene-specific translational control of GCN4 in yeast. Cell 68, 585-596.
  • 32. Feng, L., Yoon, H. & Donahue, T.F. (1994) Casein kinase II mediates multiple phosphorylation of Saccharomyces cerevisiae eIF-2α (encoded by SUI2), which is required for optimal eIF-2α function in S. cerevisiae. Mol. Cell. Biol. 14, 5139-5153.
  • 33. Yun, D.-F., Laz, T.M., Clements, J.M. & Sherman, F. (1996) mRNA sequences influencing translation and the selection of AUG initiator codons in the yeast Saccharomyces cerevisiae. Mol. Microbiol. 19, 1225-1239.
  • 34. Cigan, A.M. & Donahue, T.F. (1987) Sequences and structural features associated with translation initiator regions in yeast a review. Gene 59, 1-18.
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