Yeast topoisomerase II mutants resistant to anti-topoisomerase agents : identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide
We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperatu...
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Veröffentlicht in: | Cancer research (Chicago, Ill.) Ill.), 1994-06, Vol.54 (11), p.2943-2951 |
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creator | YA-XIA LIU YUCHU HSIUNG MEHRDAD JANNATIPOUR YEH, Y NITISS, J. L |
description | We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperature-sensitive mutation allows the isolation of recessive drug-resistant mutations. The mutagenized TOP2 gene we have used is under the control of the yeast DED1 promoter; this overexpression of TOP2 is designed to avoid isolating mutants that are drug resistant solely because the mutated topoisomerase II has low enzymatic activity. We describe three mutants that we isolated using this system. Two of the three mutants show resistance to etoposide and amsacrine, while the third mutant is partially resistant to etoposide and fluoroquinolones but not to amsacrine. DNA sequence changes have been identified in all of these mutant TOP2 genes. The mutant with partial resistance to etoposide and fluoroquinolones has an amino acid change at position 738 of TOP2, which is three amino acids from the site homologous to Ser83 of E. coli gyrA, an amino acid which had previously been shown to be an important target for resistance to quinolones in bacteria. One of the alleles that confers resistance to both etoposide and amsacrine, top2-103, has changes in amino acid 824 and amino acid 1186 of TOP2. Reconstruction of the mutations by oligonucleotide-directed mutagenesis demonstrates that the change at amino acid 824 is responsible for the drug resistance of this allele. |
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L</creator><creatorcontrib>YA-XIA LIU ; YUCHU HSIUNG ; MEHRDAD JANNATIPOUR ; YEH, Y ; NITISS, J. L</creatorcontrib><description>We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperature-sensitive mutation allows the isolation of recessive drug-resistant mutations. The mutagenized TOP2 gene we have used is under the control of the yeast DED1 promoter; this overexpression of TOP2 is designed to avoid isolating mutants that are drug resistant solely because the mutated topoisomerase II has low enzymatic activity. We describe three mutants that we isolated using this system. Two of the three mutants show resistance to etoposide and amsacrine, while the third mutant is partially resistant to etoposide and fluoroquinolones but not to amsacrine. DNA sequence changes have been identified in all of these mutant TOP2 genes. The mutant with partial resistance to etoposide and fluoroquinolones has an amino acid change at position 738 of TOP2, which is three amino acids from the site homologous to Ser83 of E. coli gyrA, an amino acid which had previously been shown to be an important target for resistance to quinolones in bacteria. One of the alleles that confers resistance to both etoposide and amsacrine, top2-103, has changes in amino acid 824 and amino acid 1186 of TOP2. Reconstruction of the mutations by oligonucleotide-directed mutagenesis demonstrates that the change at amino acid 824 is responsible for the drug resistance of this allele.</description><identifier>ISSN: 0008-5472</identifier><identifier>EISSN: 1538-7445</identifier><identifier>PMID: 8187080</identifier><identifier>CODEN: CNREA8</identifier><language>eng</language><publisher>Philadelphia, PA: American Association for Cancer Research</publisher><subject>Amino Acid Sequence ; Amsacrine - pharmacology ; Antineoplastic agents ; Base Sequence ; Biological and medical sciences ; DNA Topoisomerases, Type II - drug effects ; DNA Topoisomerases, Type II - genetics ; DNA Topoisomerases, Type II - isolation & purification ; DNA, Fungal - chemistry ; Drug Resistance - genetics ; Etoposide - pharmacology ; General aspects ; Medical sciences ; Molecular Sequence Data ; Mutation - genetics ; Pharmacology. Drug treatments ; Saccharomyces cerevisiae ; Sequence Analysis, DNA ; Yeasts - enzymology ; Yeasts - genetics ; Yeasts - isolation & purification</subject><ispartof>Cancer research (Chicago, Ill.), 1994-06, Vol.54 (11), p.2943-2951</ispartof><rights>1994 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=4108229$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8187080$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>YA-XIA LIU</creatorcontrib><creatorcontrib>YUCHU HSIUNG</creatorcontrib><creatorcontrib>MEHRDAD JANNATIPOUR</creatorcontrib><creatorcontrib>YEH, Y</creatorcontrib><creatorcontrib>NITISS, J. L</creatorcontrib><title>Yeast topoisomerase II mutants resistant to anti-topoisomerase agents : identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide</title><title>Cancer research (Chicago, Ill.)</title><addtitle>Cancer Res</addtitle><description>We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperature-sensitive mutation allows the isolation of recessive drug-resistant mutations. The mutagenized TOP2 gene we have used is under the control of the yeast DED1 promoter; this overexpression of TOP2 is designed to avoid isolating mutants that are drug resistant solely because the mutated topoisomerase II has low enzymatic activity. We describe three mutants that we isolated using this system. Two of the three mutants show resistance to etoposide and amsacrine, while the third mutant is partially resistant to etoposide and fluoroquinolones but not to amsacrine. DNA sequence changes have been identified in all of these mutant TOP2 genes. The mutant with partial resistance to etoposide and fluoroquinolones has an amino acid change at position 738 of TOP2, which is three amino acids from the site homologous to Ser83 of E. coli gyrA, an amino acid which had previously been shown to be an important target for resistance to quinolones in bacteria. One of the alleles that confers resistance to both etoposide and amsacrine, top2-103, has changes in amino acid 824 and amino acid 1186 of TOP2. Reconstruction of the mutations by oligonucleotide-directed mutagenesis demonstrates that the change at amino acid 824 is responsible for the drug resistance of this allele.</description><subject>Amino Acid Sequence</subject><subject>Amsacrine - pharmacology</subject><subject>Antineoplastic agents</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>DNA Topoisomerases, Type II - drug effects</subject><subject>DNA Topoisomerases, Type II - genetics</subject><subject>DNA Topoisomerases, Type II - isolation & purification</subject><subject>DNA, Fungal - chemistry</subject><subject>Drug Resistance - genetics</subject><subject>Etoposide - pharmacology</subject><subject>General aspects</subject><subject>Medical sciences</subject><subject>Molecular Sequence Data</subject><subject>Mutation - genetics</subject><subject>Pharmacology. Drug treatments</subject><subject>Saccharomyces cerevisiae</subject><subject>Sequence Analysis, DNA</subject><subject>Yeasts - enzymology</subject><subject>Yeasts - genetics</subject><subject>Yeasts - isolation & purification</subject><issn>0008-5472</issn><issn>1538-7445</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1994</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kM1KxDAQx4Mo67r6CEIO4q2QNEmbepPFj4UFL3rwVNJm4kbaZk1SRB_M5zNllwUvnubrN_OfmSM0p4LJrORcHKM5IURmgpf5KToL4T2FghIxQzNJZUkkmaOfV1Ah4ui2zgbXg1cB8GqF-zGqIQbsIdgwuQnBydjsL6reYMJusNXJsca2Klo3JFTjdqO8aiN4-71LOoMH-MRf_0sG6CB1aWycP-i3MC0AU1NIUufoxKguwMXeLtDL_d3z8jFbPz2slrfrbMMIiZkR1LRKaqYbxkROGGjKytxQzpqiYjmpWMEJgBGy0HnFDdXGMNqYquSEG8YW6Ho3d-vdxwgh1r0NLXSdGsCNoaaFKKqCFAm83INj04Out972yn_V-0en-tW-rkKrOuPTSTYcME6JzPOK_QLfZIzB</recordid><startdate>19940601</startdate><enddate>19940601</enddate><creator>YA-XIA LIU</creator><creator>YUCHU HSIUNG</creator><creator>MEHRDAD JANNATIPOUR</creator><creator>YEH, Y</creator><creator>NITISS, J. L</creator><general>American Association for Cancer Research</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>8FD</scope><scope>FR3</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope></search><sort><creationdate>19940601</creationdate><title>Yeast topoisomerase II mutants resistant to anti-topoisomerase agents : identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide</title><author>YA-XIA LIU ; YUCHU HSIUNG ; MEHRDAD JANNATIPOUR ; YEH, Y ; NITISS, J. L</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-h300t-f51fca8d3db335203ed1372f143b6932093640eef586d294f1dff31bf97404f33</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1994</creationdate><topic>Amino Acid Sequence</topic><topic>Amsacrine - pharmacology</topic><topic>Antineoplastic agents</topic><topic>Base Sequence</topic><topic>Biological and medical sciences</topic><topic>DNA Topoisomerases, Type II - drug effects</topic><topic>DNA Topoisomerases, Type II - genetics</topic><topic>DNA Topoisomerases, Type II - isolation & purification</topic><topic>DNA, Fungal - chemistry</topic><topic>Drug Resistance - genetics</topic><topic>Etoposide - pharmacology</topic><topic>General aspects</topic><topic>Medical sciences</topic><topic>Molecular Sequence Data</topic><topic>Mutation - genetics</topic><topic>Pharmacology. Drug treatments</topic><topic>Saccharomyces cerevisiae</topic><topic>Sequence Analysis, DNA</topic><topic>Yeasts - enzymology</topic><topic>Yeasts - genetics</topic><topic>Yeasts - isolation & purification</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>YA-XIA LIU</creatorcontrib><creatorcontrib>YUCHU HSIUNG</creatorcontrib><creatorcontrib>MEHRDAD JANNATIPOUR</creatorcontrib><creatorcontrib>YEH, Y</creatorcontrib><creatorcontrib>NITISS, J. 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L</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Yeast topoisomerase II mutants resistant to anti-topoisomerase agents : identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide</atitle><jtitle>Cancer research (Chicago, Ill.)</jtitle><addtitle>Cancer Res</addtitle><date>1994-06-01</date><risdate>1994</risdate><volume>54</volume><issue>11</issue><spage>2943</spage><epage>2951</epage><pages>2943-2951</pages><issn>0008-5472</issn><eissn>1538-7445</eissn><coden>CNREA8</coden><abstract>We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperature-sensitive mutation allows the isolation of recessive drug-resistant mutations. The mutagenized TOP2 gene we have used is under the control of the yeast DED1 promoter; this overexpression of TOP2 is designed to avoid isolating mutants that are drug resistant solely because the mutated topoisomerase II has low enzymatic activity. We describe three mutants that we isolated using this system. Two of the three mutants show resistance to etoposide and amsacrine, while the third mutant is partially resistant to etoposide and fluoroquinolones but not to amsacrine. DNA sequence changes have been identified in all of these mutant TOP2 genes. The mutant with partial resistance to etoposide and fluoroquinolones has an amino acid change at position 738 of TOP2, which is three amino acids from the site homologous to Ser83 of E. coli gyrA, an amino acid which had previously been shown to be an important target for resistance to quinolones in bacteria. One of the alleles that confers resistance to both etoposide and amsacrine, top2-103, has changes in amino acid 824 and amino acid 1186 of TOP2. Reconstruction of the mutations by oligonucleotide-directed mutagenesis demonstrates that the change at amino acid 824 is responsible for the drug resistance of this allele.</abstract><cop>Philadelphia, PA</cop><pub>American Association for Cancer Research</pub><pmid>8187080</pmid><tpages>9</tpages></addata></record> |
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subjects | Amino Acid Sequence Amsacrine - pharmacology Antineoplastic agents Base Sequence Biological and medical sciences DNA Topoisomerases, Type II - drug effects DNA Topoisomerases, Type II - genetics DNA Topoisomerases, Type II - isolation & purification DNA, Fungal - chemistry Drug Resistance - genetics Etoposide - pharmacology General aspects Medical sciences Molecular Sequence Data Mutation - genetics Pharmacology. Drug treatments Saccharomyces cerevisiae Sequence Analysis, DNA Yeasts - enzymology Yeasts - genetics Yeasts - isolation & purification |
title | Yeast topoisomerase II mutants resistant to anti-topoisomerase agents : identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide |
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