Abstract
Several pleiotropic drug sensitivities have been described in yeast. Some involve the loss of putative drug efflux pumps analogous to mammalian P-glycoproteins, others are caused by defects in sterol synthesis resulting in higher plasma membrane permeability. We have constructed a Saccharomyces cerevisiae strain that exhibits a strong crystal violet-sensitive phenotype. By selecting cells of the supersensitive strain for normal sensitivity after transformation with a wild-type yeast genomic library, a complementing 10-kb DNA fragment was isolated, a 3.4-kb subfragment of which was sufficient for complementation. DNA sequence analysis revealed that the complementing fragment comprised the recently sequenced SGE1 gene, a partial multicopy suppressor of gal11 mutations. The supersensitive strain was found to be a sge1 null mutant. Overexpression of SGE1 on a high-copy-number plasmid increased the resistance of the supersensitive strain. Disruption of SGE1 in a wild-type strain increased the sensitivity of the strain. These features of the SGE1 phenotype, as well as sequence homologies of SGE1 at the amino acid level, confirm that the Sge1 protein is a member of the drug-resistance protein family within the major facilitator superfamily (MFS).
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Amakasu H, Suzuki Y, Nishizawa M, Fukasawa T (1993) Isolation and characterization of SGE1: a yeast gene that partially suppresses the gal11 mutation in multiple copies. Genetics 134:675–683
Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (1990) Current protocols in molecular biology. John Wiley & Sons, New York, NY
Bard M, Lees ND, Burrows LS, Kleinhans FW (1978) Differences in crystal violet uptake and cation-induced death among yeast sterol mutants. J Bacteriol 135:1146–1148
Bennetzen JL, Hall BD (1982) Codon selection in yeast. J Biol Chem 257:3026–3031
Bilofsky HS, Burks C (1988) The GenBank genetic sequence data bank. Nucleic Acids Res 16:1861–1863
Chen WN, Capieaux E, Balzi E, Goffeau A (1991) The YGL022 gene encodes a putative transport protein. Yeast 7:305–308
Coque JJ, Liras P, Martin JF (1993) Genes for a β-lactamase, a penicillin-binding protein and a transmembrane protein are clustered with the cephamycin biosynthetic genes in Nocardia lactamdurans. EMBO J 12:631–639
De-Nobel JG, Barnett JA (1991) Passage of molecules through yeast cell walls: a brief essay-review. Yeast 7:313–323
De-Nobel JG, Klis FM, Munnik T, Priem J, van-den-Ende H (1990) An assay of relative cell wall porosity in Saccharomyces cerevisiae, Kluyveromyces lactis and Schizosaccharomyces pombe. Yeast 6:483–490
Fassler JS, Winston F (1989) The Saccharomyces cerevisiae SPT13/GAL11 gene has both positive and negative regulatory roles in transcription. Mol Cell Biol 9:5602–5609
Felix H (1982) Permeabilized cells. Anal Biochem 120:211–234
Goffeau A, Nakai K, Slorninski P, Risler JL (1993) The membrane proteins encoded by yeast chromosome III genes. FEBS Lett 325:112–117
Gottesman MM, Pastan I (1988) The multidrug transporter, a double-edged sword. J Biol Chem 263:12163–12166
Graham TR, Scott PA, Emr SD (1993) Brefeldin A reversibly blocks early but not late protein transport steps in the yeast secretory pathway. EMBO J 12:869–877
Guthrie C, Fink GR (1991) Guide to yeast genetics and molecular biology. Academic Press, San Diego
Haase E, Servos J, Brendel M (1992) Isolation and characterization of additional genes influencing resistance to various mutagens in the yeast Saccharomyces cerevisiae. Curr Genet 21:319–324
Hill JE, Myers AM, Koerner TJ, Tzagoloff A (1986) Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast 2:163–167
Hoffman CS, Winston F (1987) A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene 57:267–272
Klebe RJ, Harriss JV, Sharp ZD, Douglas MG (1983) A general method for polyethylene-glycol-induced genetic transformation of bacteria and yeast. Gene 25:333–341
Kuchler K, Sterne RE, Thorner J (1989) Saccharomyces cerevisiae STE6 gene product: a novel pathway for protein export in eukaryotic cells. EMBO J 8:3973–3984
Kyte J, Doolittle RF (1982) A simple method for displaying the hydropathic character of a protein. J Mol Biol 157:105–132
Lehninger AL (1977) Biochemistry. Worth Publishers, New York
Leppert G, McDevitt R, Falco SC, Van-Dyk TK, Ficke MB, Golin J (1990) Cloning by gene amplification of two loci conferring multiple drug resistance in Saccharomyces. Genetics 125:13–20
Marger MD, Saier M Jr (1993) A major superfamily of transmembrane facilitators that catalyse uniport, symport and antiport. Trends Biochem Sci 18:13–20
McCammon MT, Hartmann MA, Bottema CD, Parks LW (1984) Sterol methylation in Saccharomyces cerevisiae. J Bacteriol 157:475–483
McGrath JP, Varshavsky A (1989) The yeast STE6 gene encodes a homologue of the mammalian multidrug resistance P-glycoprotein. Nature 340:400–404
Meyers S, Schauer W, Balzi E, Wagner M, Goffeau A, Golin J (1992) Interaction of the yeast pleiotropic drug resistance genes PDR1 and PDR5. Curr Genet 21:431–436
Morita T, Yanagihara Y (1985) Osmotic-sensitive mutants of Saccharomyces cerevisiae as screening organisms for promutagens and procarcinogens. Chem Pharm Bull Tokyo 33:1576–1582
Nishizawa M, Suzuki Y, Nogi Y, Matsumoto K, Fukasawa T (1990) Yeast Gal11 protein mediates the transcriptional activation signal of two different transacting factors, Gal4 and general regulatory factor I/repressor/activator site binding protein 1/translation upstream factor. Proc Natl Acad Sci USA 87:5373–5377
Nitiss J, Wang JC (1988) DNA topoisomerase-targeting antitumor drugs can be studied in yeast. Proc Natl Acad Sci USA 85:7501–7505
Peden KW (1983) Revised sequence of the tetracycline-resistance gene of pBR322. Gene 22:277–280
Rank GH, Robertson A, Phillips K (1975) Reduced plasma membrane permeability in a multiple cross-resistant strain of Saccharomyces cerevisiae. J Bacteriol 122:359–366
Riordan JR, Rommens JM, Kerem B, Alon N, Rozmahel R, Grzelczak Z, Zielenski J, Lok S, Plavsic N, Chou JL (1989) Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science 245:1066–1073
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Sauer B (1992) Identification of cryptic lox sites in the yeast genome by selection for Cre-mediated chromosome translocations that confer multiple drug resistance. J Mol Biol 223:911–928
Scherrer R, Louden L, Gerhardt P (1974) Porosity of the yeast cell wall and membrane. J Bacteriol 118:534–540
Sengstag C (1993) The sequence of Saccharomyces cerevisiae cloning vector pCS19 following direct selection for DNA inserts. Gene 124:141–142
Sengstag C, Hinnen A (1987) The sequence of the Saccharomyces cerevisiae gene PH02 codes for a regulatory protein with unusual amino acid composition. Nucleic Acids Res 15:233–246
Servos J, Haase E, Brendel M (1993) Gene SNQ2 of Saccharomyces cerevisiae, which confers resistance to 4-nitroquinoline-N-oxide and other chemicals, encodes a 169 kDa protein homologous to ATP-dependent permeases. Mol Gen Genet 236:214–218
Sherman F (1991) Getting started with yeast. Methods Enzymol 194:3–21
Sikorski RS, Hieter P (1989) A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122:19–27
Suzuki Y, Nogi Y, Abe A, Fukasawa T (1988) GAL11 protein, an auxiliary transcription activator for genes encoding galactose-metabolizing enzymes in Saccharomyces cerevisiae. Mol Cell Biol 8:4991–4999
Ulaszewski S, Balzi E, Goffeau A (1987) Genetic and molecular mapping of the pma1 mutation conferring vanadate resistance to the plasma membrane ATPase from Saccharomyces cerevisiae. Mol Gen Genet 207:38–46
Wang M, Balzi E, Van Dyck L, Golin J, Goffeau A (1992) Sequencing of the yeast multidrug resistance PDR5 gene encoding a putative pump for drug efflux. Yeast 8:S528
Winsor B, Potter AA, Karst F, Nestmann ER, Lacroute F (1987) Characterization of a yeast mutation ise1 that enhances permeability of Saccharomyces cerevisiae. Environ Mutagen 9:114
Zimmermann FK, Kern R, Rasenberger H (1975) A yeast strain for simultaneous detection of induced mitotic crossing over, mitotic gene conversion and reverse mutation. Mutat Res 28:381–388
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Communicated by C. P. Hollenberg
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Ehrenhofer-Murray, A.E., Würgler, F.E. & Sengstag, C. The Saccharomyces cerevisiae SGE1 gene product: a novel drug-resistance protein within the major facilitator superfamily. Molec. Gen. Genet. 244, 287–294 (1994). https://doi.org/10.1007/BF00285456
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DOI: https://doi.org/10.1007/BF00285456