Summary
The cryptopleurine resistance gene, cry1, of Saccharomyces cerevisiae has been molecularly cloned using genetic complementation of cryptopleurine sensitivity by the cryptopleurine resistance gene contained in a clone library prepared from DNA of a cryptopleurine resistant strain. Analysis of RNA transcripts indicated that the cry1 gene is the template for a transcript of approximately 900 bases and that the primary transcript contains an intron of approximately 300 bases. In vitro hybrid selection translation experiments indicated that this transcript encodes a protein of molecular weight 17 kilodaltons which on two-dimensional SDS polyacrylamide gels exactly coincides with ribosomal protein rp59. Further analysis showed that when the gene was present on a plasmid of about five copies per cell the amount of messenger RNA was elevated approximately five-fold compared to a cell that had only a single chromosomal copy. The rate of synthesis of ribosomal protein rp59 was not detectably elevated. These data suggest that the cry1 gene is regulated, at least in part, post-transcriptionally.
Similar content being viewed by others
References
Branlant C, Krol A, Machatt A, Ebel J-P (1981) The secondary structure of the protein L1 binding region of ribosomal 23S RNA. Homologies with putative secondary structure of the L11 mRNA and of a region of mitochondrial 16s rRNA. Nucl Acids Res 9:293–307
Bromley S, Hereford L, Rosbash M (1982) Further evidence that the rna2 mutation of Saccharomyces cerevisiae affects mRNA processing. Mol Cell Biol 2:1205–1211
Davis RW, Thomas M, Cameron J, St. John TP, Scherer S, Padgett RA (1980) Rapid DNA isolations for enzymatic and hybridization analysis. Methods Enzymol 65:404–411
Dennis PP, Fiil NP (1979) Transcriptional and post-transcriptional control of RNA polymerase and ribosomal protein genes cloned on composite ColE1 plasmids in the bacterium Escherichia coli. J Biol Chem 254:7540–7547
Dennis PP, Nomura M (1974) Stringent control of ribosomal protein gene expression in Escherichia coli. Proc Natl Acad Sci USA 71:3819–3823
Erhart E, Hollenberg CP (1983) The presence of a defective LEU2 gene on 2μ DNA recombinant plasmids of Saccharomyces cerevisiae is responsible for curing and high copy number. J Bacteriol 156:625–635
Fallon AM, Jinks CS, Strycharz GD, Nomura M (1979) Regulation of ribosomal protein synthesis in Escherichia coli by selective mRNA inactivation. Proc Natl Acad Sci USA 76:3411–3415
Fiil NP, Bendiak D, Collins J, Friesen JD (1979) Expression of Escherichia coli ribosomal protein and RNA polymerase genes cloned on plasdmids. Mol Gen Genet 173:39–50
Fried HM, Warner JR (1981) Cloning of yeast gene for trichodermin resistance and ribosomal protein L3. Proc Natl Acad Sci USA 78:238–242
Fried HM, Warner JR (1982) Molecular cloning and analysis of yeast gene for cycloheximide resistance and ribosomal protein L29. Nucl Acids Res 10:3133–3148
Friesen JD, Fiil NP, Dennis PP, Downing WL, An G, Holowachuk E (1980) Biosynthetic regulation of rp1J, rp1L, rpoB and rpoC in Escherichia coli. In: Chambliss G, Craven GR, Davies J, Davis K, Kahan L, Nomura M (eds) Ribosomes, structure, function and genetics. University Park Press, Baltimore, Maryland, pp 719–742
Friesen JD, Tropak M, An G (1983) Mutations in the rp1J leader of Escherichia coli that abolish feedback regulation. Cell 32:361–369
Gorenstein C, Warner JR (1976) Coordinate regulation of the synthesis of eukaryotic ribosomal proteins. Proc Natl Acad Sci USA 73:1547–1551
Gourse RL, Thurlow DL, Gerbi SA, Zimmerman RA (1981) Specific binding of a prokaryotic ribosomal protein to a eukaryotic ribosomal RNA: Implications for evolution and autoregulation. Proc Natl Acad Sci USA 78:2722–2726
Grant P, Sanchez L, Jimenez A (1974) Cryptopleurine resistance: Genetic locus for a 40S ribosomal component in Saccharomyces cerevisiae. J Bacteriol 120:1308–1314
Guarente L, Mason T (1983) Heme regulates transcription of the CYC1 gene of S. cerevisiae via an upstream activation site. Cell 32:1279–1286
Hartwell LC, McLaughlin C, Warner JR (1970) Identification of ten genes that control ribosome formation in yeast. Mol Gen Genet 109:42–56
Kief DR, Warner JR (1981) Coordinate control of syntheses of ribosomal ribonucleic acid and ribosomal proteins during nutritional shift-up in Saccharomyces cerevisiae. Mol Cell Biol 1:1007–1015
Kim CH, Warner JR (1983) Mild temperature shock alters the transcription of a discrete class of Saccharomyces cerevisiae genes. Mol Cell Biol 3:457–465
Larkin JC, Woolford JL Jr (1983) Molecular cloning and analysis of the CRY1 gene: a yeast ribosomal protein gene. Nucl Acids Res 11:403–420
Lindahl L, Zengel JM (1982) Expression of ribosomal genes in bacteria. Adv Genet 21:53–121
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning. A Laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
McNeil JB, Friesen JD (1981) Expression of the Herpes simplex virus thymidine kinase gene in Saccharomyces cerevisiae. Mol Gen Genet 184:386–393
Meade JH, Riley MI, Manney TR (1977) Expression of cryptopleurine resistance in Saccharomyces cerevisiae. J Bacteriol 129:1428–1434
Mortimer RK, Schild D (1980) Genetic map of Saccharomyces cerevisiae. Microbiol Rev 44:519–571
Nomura M, Dean D, Yates JL (1982) Feedback regulation of ribosomal protein synthesis in Escherichia coli. Trends in Biochemical Sciences 7:92–95
Olins P, Nomura M (1981a) Translational regulation by ribosomal protein S8 in Escherichia coli: Structural homology between rRNA binding site and feedback target on mRNA. Nucl Acids Res 9:1757–1764
Olins P, Nomura M (1981b) Regulation of the S10 ribosomal protein operon in E. coli: Nucleotide sequence at the start of the operon. Cell 26:205–211
Orr-Weaver TL, Szostak JW, Rothstein RJ (1981) Yeast transformation: A model system for the study of recombination. Proc Natl Acad Sci USA 78:6354–6358
Pearson NJ, Fried HM, Warner JR (1982) Yeast use translational control to compensate for extra copies of a ribosomal protein gene. Cell 29:347–355
Pedersen S, Reeh S, Parker J, Watson RJ, Friesen JD, Fiil NP (1976) Analysis of the proteins synthesized in ultraviolet light-irradiated Escherichia coli following infection with the bacteriophage λdrifd18 and λdfus3. Mol Gen Genet 144:339–344
Rosbash M, Harris PKW, Woolford JL Jr, Teem JL (1981) The effect of temperature-sensitive RNA mutants on the transcription products from cloned ribosomal protein genes of yeast. Cell 24:679–686
Schultz LD, Friesen JD (1983) Nucleotide sequence of the tcm1 gene (ribosomal protein L3) of Saccharomyces cerevisiae. J Bacteriol 155:8–14
Skogerson L, McLaughlin C, Wakatama E (1973) Modification of ribosomes in cryptopleurine-resistant mutants of yeast. J Bacteriol 116:818–822
Storms RK, McNeil JB, Khandekar PS, An G, Parker J, Friesen JD (1979) Chimeric plasmids for cloning of deoxyribonucleic acid in S. cerevisiae. J Bacteriol 140:73
Warner J (1982) The yeast ribosome: structure, function and synthesis. In: Strathern JN, Jones EW, Broach JR (eds) The molecular biology of the yeast Saccharomyces. Metabolism and gene expression. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, pp 529–560
Author information
Authors and Affiliations
Additional information
Communicated by K. Isono
Rights and permissions
About this article
Cite this article
Himmelfarb, H.J., Vassarotti, A. & Friesen, J.D. Molecular cloning and biosynthetic regulation of the cry1 gene of Saccharomyces cerevisiae . Mol Gen Genet 195, 500–506 (1984). https://doi.org/10.1007/BF00341453
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00341453