Summary
A DNA fragment conferring resistance to zinc and cadmium ions in the yeast Saccharomyces cerevisiae was isolated from a library of yeast genomic DNA. Its nucleotide sequence revealed the presence of a single open reading frame (ORF; 1326 bp) having the potential to encode a protein of 442 amino acid residues (molecular mass of 48.3 kDa). A frameshift mutation introduced within the ORF abolished resistance to heavy metal ions, indicating the ORF is required for resistance. Therefore, we termed it the ZRC1 (zinc resistance conferring) gene. The deduced amino acid sequence of the gene product predicts a rather hydrophobic protein with six possible membrane-spanning regions. While multiple copies of the ZRC1 gene enable yeast cells to grow in the presence of 40 mM Zn2+, a level at which wild-type cells cannot survive, the disruption of the chromosomal ZRC1 locus, though not a lethal event, makes cells more sensitive to zinc ions than are wild-type cells.
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References
Adzuma K, Ogawa T, Ogawa H (1984) Primary structure of the RAD52 gene in S. cerevisiae. Mol Cell Biol 4:2735–2744
Butt TR, Ecker DJ (1987) Yeast metallothionein and applications in biotechnology. Microbiol Rev 51:351–364
Butt TR, Sternberg E, Herd J, Crooke ST (1984) Cloning and expression of a yeast copper metallothionein gene. Gene 27:23–33
Byrd J, Berger RM, McMillin DR, Wright CF, Hamer D, Winge DR (1988) Characterization of the copper-thiolate cluster in yeast metallothionein and two truncated mutants. J Biol Chem 263:6688–6694
Ecker DJ, Butt TR, Sternberg EJ, Neeper MP, Debouck C, Gorman JA, Crooke ST (1986) Yeast metallothionein function in metal detoxification. J Biol Chem 261:16895–16900
Evans RM, Hollenberg SM (1988) Zinc fingers: gilt by association. Cell 52:1–3
Feinberg AP, Vogelstein B (1983) A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 132:6–13
Fogel S, Welch JW (1982) Tandem gene amplification mediates copper resistance in yeast. Proc Natl Acad Sci USA 79:5342–5346
Fogel S, Welch JW, Karin M (1983) Gene amplification in yeast: CUP1 copy number regulates copper resistance. Curr Genet 7:1–9
Fukuda Y, Saikusa T, Watanabe K, Shimosaka M, Murata K, Kimura A (1986) Cloning of genes enhancing the resistance of Saccharomyces cerevisiae to zinc and cadmium ions. Agric Biol Chem 50:1341–1343
Furst P, Hu S, Hackett R, Hamer D (1988) Copper activates metallothionein gene transcription by altering the conformation of a specific DNA binding protein. Cell 55:705–717
Grill E, Winnacker EL, Zenk MH (1986) Synthesis of seven different homologous phytochelatins in metal exposed Schizosaccharomyces pombe cells. FEBS Lett 197:115–120
Gros P, Croop J, Housman D (1986) Mammalian multidrug resistance gene: complete eDNA sequence indicates strong homology to bacterial transport protein. Cell 47:371–380
Hartman H, Weser W (1985) Cobalt-(cystein)4 tetrahedra in yeast cobalt(II) thionein. Biochem Biophys Res Commun 132:277–283
Henikoff S (1984) Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene 28:351–359
Ikemura T (1982) Correlation between the abundance of yeast transfer RNAs and the occurrence of the respective codons in protein genes. J Mol Biol 158:573–597
Inouye S, Nakazawa A, Nakazawa T (1981) Molecular cloning of TOL genes, xylB and xylE in Escherichia coli. J Bacteriol 145:1137–1143
Ito H, Fukuda Y, Murata K, Kimura A (1983) Transformation of intact cells treated with alkali cations. J Bacteriol 153:163–168
Jones JS, Weber S, Prakash L (1988) The Saccharomyces cerevisiae RAD18 gene encodes a protein that contains potential zinc finger domains for nucleic acid binding and a putative nucleotide binding sequence. Nucleic Acids Res 16:7119–7131
Kanehisa M (1982) Los Alamos sequence analysis package for nucleic acids and proteins. Nucleic Acids Res 10:183–196
Klein P, Kanehisa M, Delisi C (1985) The detection and classification of membrane spanning proteins. Biochim Biophys Acta 815:468–475
Kondo N, Isobe M, Imai K, Goto J, Murasugi A, Hayashi Y (1983) Structure of cadystin, the unit-peptide of cadmium-binding peptides induced in a fission yeast Schizosaccharomyces pombe. Tetrahedron Lett 24:925–928
Kornfeld K, Kornfeld S (1985) Assembly of asparagine-linked oligosaccharide. Annu Rev Biochem 54:631–664
Kyte J, Doolittle RF (1982) A simple method for displaying the hydropathic character of a protein. J Mol Biol 157:105–132
Lerch K (1980) Copper metallothionein, a copper-binding protein from Neurospora crssa. Nature 284:368–370
Lipman DJ, Pearson WR (1985) Rapid and sensitive protein similarity searches. Science 227:1435–1441
Mandel M, Higa A (1970) Calcium-dependent bacteriophage DNA infection. J Mol Biol 53:159–162
Maniatis TE, Fritsch F, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY
McClanahan T, McEntee K (1984) Specific transcripts are elevated in Saccharomyces cerevisiae in response to DNA damage. Mol Cell Biol 4:2356–2363
McClanahan T, McEntee K (1986) DNA damage and heat shock dually regulate genes in Saccharomyces cerevisiae. Mol Cell Biol 6:90–96
Mehra RK, Tarbet EB, Gray WR, Winge DR (1988) Metal-specific synthesis of two metallothioneins and γ-glutamyl peptides in Candida glabrata. Proc Natl Acad Sci USA 85: 8815–8819
Murasugi A, Wada C, Hayashi Y (1985) Cadmium-binding peptide induced in fission yeast Schizosaccharomyces pombe. J Biochem 96:1561–1564
Murata K, Fukuda Y, Shimosaka M, Watanabe K, Saikusa T, Kimura A (1985) Phenotypic character of the methylglyoxal resistance gene in Saccharomyces cerevisiae: expression in E. coli and application to breeding wild-type yeast strains. Appl Environ Microbiol 50:1200–1207
Nishizawa M, Araki R, Teranishi Y (1989) Identification of an upstream activating sequence and an upstream repressible sequence of pyruvate kinase gene of the yeast Saccharomyces cerevisiae. Mol Cell Biol 9:442–451
Ostrowski J, Burdzy GJ, Kredich NM (1987) DNA sequence of the cysB regions of Salmonella typhimurium and E. coli. J Biol Chem 262:5999–6005
Perozzi G, Prakash S (1986) RAD7 gene of S. cerevisiae: transcripts, nucleotide sequence analysis, and functional relationship between the RAD7 and RAD23 gene products. Mol Cell Biol 6:1497–1507
Perron CY, Vieira J, Messing J (1985) Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mpl8 and pUC19 vectors. Gene 33:103–119
Reynolds P, Higins DR, Prakash L, Prakash S (1985a) The nucleotide sequence of the RAD3 gene of S. cerevisiae: a potential adenine nucleotide binding amino acid sequence and a nonessential acidic calboxyl terminal region. Nucleic Acids Res 13:2357–2372
Reynolds P, Weber S, Prakash L (1985b) RAD6 gene of S. cerevisiae encodes a protein containing a tract of 13 consecutive aspartates. Proc Natl Acad Sci USA 82:168–172
Rine J, Hansen W, Hardeman E, Davis RW (1983) Targeted selection of recombinant clones through gene dosage effects. Proc Natl Acad Sci USA 80:6750–6754
Rothstein R (1983) One-step gene disruption in yeast. Methods Enzymol 101:202–211
Ruby SW, Szostak JW (1985) Specific Saccharomyces cerevisiae genes are expressed in response to DNA-damage agents. Mol Cell Biol 5:75–84
Sanger F, Micklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Shatzman AR, Kosman DJ (1979) Characterization of two copperbinding components of the fungus Dactylium dendroides. Arch Biochem Biophys 194:226–235
Sherman F, Fink GR, Hicks JB (1986) Laboratory course manual for methods in yeast genetics, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY
Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 8:503–517
Struhl K, Davis RW (1981) Transcription of the HIS3 gene region in Saccharomyces cerevisiae. J Mol Biol 152:535–552
Vieira J, Messing J (1987) Production of single-stranded plasmid DNA. Methods Enzymol 153:3–11
Weiss WA, Friedberg EC (1985) Molecular cloning and characterization of the yeast RAD10 gene and expression of RAD10 protein in E. coli. EMBO J 4:1575–1582
Winge DR, Nielson RB, Gray WR, Hamer DH (1985) Yeast metallothionein sequence and metal-binding properties. J Biol Chem 260:14464–14470
Yang E, Friedberg EC (1984) Molecular cloning and nucleotide sequence analysis of the S. cerevisiae RAD1 gene. Mol Cell Biol 4:2161–2169
Zaret KS, Sherman F (1982) DNA sequence required for efficient transcription termination in yeast. Cell 28:563–573
Zukowski MM, Gaffney DF, Speck D, Kauffman A, Findeli A, Wisecup A, Lecocq JP (1983) Chromogenic identification of genetic regulatory signals in Bacillus subtilis based on expression of a cloned Pseudomonas gene. Proc Natl Acad Sci USA 80:1101–1105
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Communicated by C.P. Hollenberg
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Kamizono, A., Nishizawa, M., Teranishi, Y. et al. Identification of a gene conferring resistance to zinc and cadmium ions in the yeast Saccharomyces cerevisiae . Molec. Gen. Genet. 219, 161–167 (1989). https://doi.org/10.1007/BF00261172
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DOI: https://doi.org/10.1007/BF00261172