Summary
CYP1 is a trans acting regulatory locus modulating both iso 1- and iso 2-cytochrome c synthesis. Genetical analysis of various mutated alleles has allowed us to identify the gene product as a positive regulatory element.
The region of the target sequence of the CYP1 product on the iso 2-cytochrome c structural gene was located by molecular and genetic analysis of two cis acting mutations located at the CYP3 locus: CYP3-36 and CYP3-4, which have been shown to arise from the integration of TY1 elements near the promoter site. Determination of the amount of iso 2-cytochrome c synthesized by strains bearing various genetic constructions, in which the cis acting mutations were associated with different alleles of the CYP1 trans acting locus, showed that TY1 inserted into CYP3-36 extinguishes the activation function due to a mutated overproducer allele CYP1-18, while CYP3-4 amplifies this function. This result identifies at least a part of the target sequence of the CYP1 product within the region separating the two TY1 insertions.
To clone the CYP1 gene, we took advantage of the iso 2-cytochrome c overproducer phenotype of the mutated allele CYP1-18, which confers a Lactate+ phenotype on an iso 1-cytochrome c-deficient strain. Such a phenotype allowed the isolation of a recombinant plasmid YEpJFM1 carrying the mutated allele, able to complement on lactate medium a lactate - recipient strain. The identity of the YEpJFM1 sequence with the chromosomal gene was confirmed by homologous recombination at the CYP1 locus.
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References
Bach ML, Lacroute F, Botstein D (1979) Evidence for transcriptional regulation of orotidine-5′-phosphate decarboxylase in yeast by hybridization of mRNA to the yeast structural gene cloned in E. coli. Proc Natl Acad Sci USA 76:386–390
Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmids. Nucleic Acids Res 7:1513–1523
Blane H, Gerbaud C, Slonimski PP, Guerineau M (1979) Stable yeast transformation with chimeric plasmids using a 2 micron circular DNA-less strain as a recipient. Mol Gen Genet 176:335–342
Boyer HW, Roulland-Dussois D (1969) A complementation analysis of the restriction and modification of DNA Escherichia coli. J Mol Biol 41:459–472
Claisse M, Péré-Aubert G, Clavilier L, Slonimski PP (1970) Méthode d'estimation de la concentration des cytochromes dans les cellules entières de levure. Eur J Biochem 16:430–438
Clavilier L, Péré G, Slonimski PP, Somlo M (1964) Contrôle génétique de la biosynthèse des isocytochromes c. The Sixth International Congress of Biochemistry, New York, pp 673–674
Clavilier L, Péré G, Slonimski PP (1969) Mise en évidence de plusieurs loci indépendants impliqués dans la synthèse de l'iso 2-cytochrome c chez la levure. Mol Gen Genet 104:195–218
Clavilier L, Péré-Aubert G, Somlo M, Slonimski PP (1976) Réseau d'interactions entre des gènes non liés: régulation synergique ou antagoniste de la synthèse de l'iso l-cytochrome c, de l'iso 2-cytochrome c, et du cytochrome b2. Biochimie 58:155–172
Covarrubias L, Bolivar F (1982) Construction and characterization of new cloning vehicles. VI. Plasmid pBR329, a new derivative of pBR328 lacking the 482 base pair inverted duplication. Gene 17:79–89
Downie JA, Stewart JW, Brockman N, Schweingurber AM, Sherman F (1977) The structural gene for iso 2-cytochrome c. J Mol Biol 113:369–384
Dretzen G, Bellard M, Sassone-Corsi P, Chambon P (1981) A reliable method for the recovery of DNA fragments from agarose and acrylamide gels. Anal Biochem 112:295–298
Errede B, Cardillo TS, Sherman F, Dubois E, Deschamps J, Wiame JM (1980) Mating signals control expression of mutations resulting from insertion of a transposable repetitive element adjacent to diverse yeast genes. Cell 22:427–436
Gerbaud C, Fournier P, Blanc H, Aigle M, Heslot H, Guerineau M (1979) High frequency of yeast transformation by plasmids carrying part or entire 2 micron yeast plasmid. Gene 5:233–253
Gerbaud C, Elmerich C, Tandeau de Marsac N, Chocat P, Charpin N, Guerineau M, Aubert JP (1981) Construction of new yeast vectors and cloning of the Nif (Nitrogen fixation) gene cluster of Klebsiella pneumoniae in yeast. Curr Genet 3:173–180
Cuarente L, Mason T (1983) Heme regulates transcription of the CYC1 gene of S. cerevisiae via an upstream activation site. Cell 32:1279–1286
Guarente L, Lalonde B, Gifford P, Alani E (1984) Distinctly regulated tandem upstream activation site mediate catabolite repression of the CYC1 gene of S. cerevisiae. Cell 36:503–511
Guerry P, Leblanc DJ, Falkow S (1973) General method for the isolation of plasmid deoxyribonucleic acid. J Bacteriol 116:1064–1066
Hawthorne DC (1969) The selection of nonsense suppressors in yeast. Mutat Res 7:187–197
Hereford L, Fahrner K, Woolford J Jr, Rosbash M, Kaback DB (1979) Isolation of yeast histone genes H2A and H2B. Cell 18:1261–1271
Hicks JB, Hinnen A, Fink GR (1978) Properties of yeast transformation. Cold Spring Harbor Symp Quant Biol 43:1305–1313
Hinnen A, Hicks JB, Fink GR (1978) Transformation of yeast. Proc Natl Acad Sci USA 75:1929–1933
Hohn B (1979) In vitro packaging of lambda and cosmid DNA. Methods Enzymol 68:299–309
Hohn B, Collins J (1980) A small cosmid for efficient cloning of large DNA fragments. Gene 11:291–298
Ito H, Fukuda Y, Murata K, Kimma A (1983) Transformation of intact yeast cells treated with alkali cations. J Bacteriol 153:163–168
Lemontt JF (1977) Pathways of ultraviolet mutability in S. cerevisiae. III. Genetic analysis and properties of mutants resistant to ultraviolet-induced forward mutations. Mutat Res 43:179–204
Lemontt JF, Fugit DR, Mackay VL (1980) Pleiotropic mutations at the tup1 locus that affect the expression of mating type dependent functions in S. cerevisiae. Genetics 94:899–920
Mandel M, Higa A (1970) Calcium-dependent bacteriophage DNA infection. J Mol Biol 53:159–162
Maniatis T, Fritsch EF, Sambrook J (1982) In: Cold Spring Harbor Laboratory (ed) Molecular cloning. A laboratory manual. Cold Spring Harbor, New York
McKnight GL, Cardillo TS, Sherman F (1981) An extensive deletion causing overproduction of yeast iso 2-cytochrome c. Cell 25:409–419
Miller JH (1972) In: Cold Spring Harbor Laboratory (ed) Experiments in molecular genetics. Cold Spring Harbor, New York
Montgomery DL, Boss JM, McAndrew SJ, Marr L, Walthall DA, Zitomer RS (1982) The molecular characterization of three transcriptional mutations in the yeast iso 2-cytochrome c gene. J Biol Chem 257:7756–7761
Mortimer RK, Hawthorne DC (1966) Genetic mapping in S. cerevisiae. Genetics 53:165–173
Muller MJ (1932) Further studies on the nature and cause of gene mutations. Proc 6th Int Congr Genet 1:213–255
Nasmyth KA, Reed SI (1980) Isolation of genes by complementation in yeast: molecular cloning of a cell-cycle gene. Proc Natl Acad Sci USA 77:2119–2123
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
Petrochilo E, Verdière J (1977) A glycine to serine substitution identifies the CYP3 locus as the structural gene of iso 2-cytochrome c in Saccharomyces cerevisiae. Biochem Biophys Res Commun 79:364–371
Rothstein RJ, Sherman F (1980) Genes affecting the expression of cytochrome c in yeast: genetic mapping and genetic interactions. Genetics 94:871–889
Sels AA, Fukuhara H, Péré G, Slonimski PP (1965) Cinétique de la biosynthèse induite de l'iso l-cytochrome c et de l'iso 2-cytochrome c au cours de l'adaptation à l'oxygène. Biochim Biophys Acta 95:486–502
Sherman F, Stewart JW, Parker J, Inhaber E, Shipman NA, Putterman GJ, Gardisky RL, Margoliash E (1968) The mutational alteration of the primary structure of yeast iso 1-cytochrome c. J Biol Chem 243:5446–5456
Sherman F, Stewart JW, Helms C, Downie JA (1978) Chromosome mapping of the CYC7 gene determining yeast iso 2-cytochrome c. Structural and regulatory regions. Proc Natl Acad Sci USA 75:1437–1441
Slonimski PP (1953) Formation des enzymes respiratoires chez la levure. Masson et Cie, Paris
Slonimski PP, Acher R, Péré G, Sels AA, Somlo M (1963) Eléments du système respiratoire et leur régulation: cytochrome et isocytochromes. In: Intern symp mechanisms of regulation of cellular activities in microorganisms. CNRS, Marseille, Gordon and Breach, pp 435–461
Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517
Stark HC, Fugit D, Mowshowitz DB (1980) Pleiotropic properties of a mutant of yeast insensitive to catabolite repression. Genetics 94:921–928
Verdière J, Petrochilo E (1975) CYP3, a likely candidate for the structural gene of iso 2-cytochrome c in S. cerevisiae. Biochem Biophys Res Commun 67:1451–1458
Verdière J, Petrochilo E (1979) CYP3-15: an up-promoter mutation at the iso 2-cytochrome c structural gene locus and its interaction with other independent regulatory mutations of cytochrome c synthesis in Saccharomyces cerevisiae. Mol Gen Genet 175:209–216
Wickner RB (1974) Mutants of S. cerevisiae that incorporate deoxythymidine-5-monophosphate into DNA in vivo. J Bacteriol 117:252–260
Williamson VH (1983) Transposable elements in yeast. Int Rev Cytol 83:1–25
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Communicated by W. Gajewski
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Verdière, J., Creusot, F. & Guérineau, M. Regulation of the expression of iso 2-cytochrome c gene in S. cerevisiae: cloning of the positive regulatory gene CYP1 and identification of the region of its target sequence on the structural gene CYP3 . Mol Gen Genet 199, 524–533 (1985). https://doi.org/10.1007/BF00330769
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DOI: https://doi.org/10.1007/BF00330769