Identification of protein complexes required for efficient sister chromatid cohesion
Ctf8p is a component of Ctf18-RFC, an alternative replication factor C-like complex required for efficient sister chromatid cohesion in Saccharomyces cerevisiae. We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes r...
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| Veröffentlicht in: | Molecular biology of the cell 2004-04, Vol.15 (4), p.1736-1745 |
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| container_title | Molecular biology of the cell |
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| creator | Mayer, Melanie L Pot, Isabelle Chang, Michael Xu, Hong Aneliunas, Victoria Kwok, Teresa Newitt, Rick Aebersold, Ruedi Boone, Charles Brown, Grant W Hieter, Philip |
| description | Ctf8p is a component of Ctf18-RFC, an alternative replication factor C-like complex required for efficient sister chromatid cohesion in Saccharomyces cerevisiae. We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes required for efficient sister chromatid cohesion. We then assessed proficiency of cohesion at three chromosomal loci in strains containing deletions of the genes identified in the ctf8 SGA screen. Deletion of seven genes (CHL1, CSM3, BIM1, KAR3, TOF1, CTF4, and VIK1) resulted in defective sister chromatid cohesion. Mass spectrometric analysis of immunoprecipitated complexes identified a physical association between Kar3p and Vik1p and an interaction between Csm3p and Tof1p that we confirmed by coimmunoprecipitation from cell extracts. These data indicate that synthetic genetic array analysis coupled with specific secondary screens can effectively identify protein complexes functionally related to a reference gene. Furthermore, we find that genes involved in mitotic spindle integrity and positioning have a previously unrecognized role in sister chromatid cohesion. |
| doi_str_mv | 10.1091/mbc.E03-08-0619 |
| format | Article |
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We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes required for efficient sister chromatid cohesion. We then assessed proficiency of cohesion at three chromosomal loci in strains containing deletions of the genes identified in the ctf8 SGA screen. Deletion of seven genes (CHL1, CSM3, BIM1, KAR3, TOF1, CTF4, and VIK1) resulted in defective sister chromatid cohesion. Mass spectrometric analysis of immunoprecipitated complexes identified a physical association between Kar3p and Vik1p and an interaction between Csm3p and Tof1p that we confirmed by coimmunoprecipitation from cell extracts. These data indicate that synthetic genetic array analysis coupled with specific secondary screens can effectively identify protein complexes functionally related to a reference gene. 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We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes required for efficient sister chromatid cohesion. We then assessed proficiency of cohesion at three chromosomal loci in strains containing deletions of the genes identified in the ctf8 SGA screen. Deletion of seven genes (CHL1, CSM3, BIM1, KAR3, TOF1, CTF4, and VIK1) resulted in defective sister chromatid cohesion. Mass spectrometric analysis of immunoprecipitated complexes identified a physical association between Kar3p and Vik1p and an interaction between Csm3p and Tof1p that we confirmed by coimmunoprecipitation from cell extracts. These data indicate that synthetic genetic array analysis coupled with specific secondary screens can effectively identify protein complexes functionally related to a reference gene. Furthermore, we find that genes involved in mitotic spindle integrity and positioning have a previously unrecognized role in sister chromatid cohesion.</description><subject>Chromatids - ultrastructure</subject><subject>Chromosomes, Fungal</subject><subject>Epitopes - chemistry</subject><subject>Fungal Proteins - physiology</subject><subject>Gene Deletion</subject><subject>Genetic Techniques</subject><subject>Genome, Fungal</subject><subject>Green Fluorescent Proteins</subject><subject>Luminescent Proteins - metabolism</subject><subject>Mass Spectrometry</subject><subject>Mutation</subject><subject>Oligonucleotide Array Sequence Analysis</subject><subject>Precipitin Tests</subject><subject>Saccharomyces cerevisiae</subject><subject>Saccharomyces cerevisiae - physiology</subject><subject>Saccharomyces cerevisiae Proteins - physiology</subject><issn>1059-1524</issn><issn>1939-4586</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkTtPwzAURi0EorxmNpSJLdQ3dux6YEBVeUiVWLpbiXNNjZI4tVME_x6jVjwmJtvyOVff1UfIJdAboAqmXW1uFpTldJZTAeqAnIBiKuflTBymOy1VDmXBJ-Q0xldKgXMhj8kEuOSFBH5CVk8N9qOzzlSj833mbTYEP6LrM-O7ocV3jFnAzdYFbDLrQ4Y2wS5JWXRxxJCZdfBdsptkrDGmKefkyFZtxIv9eUZW94vV_DFfPj88ze-WuSkFH3NAngKCEjU3zFqDDbOmok1ditoIWaMUdYMFygawtkIyVjLKQHBpQaXXGbndjR22dYeNSZlC1eohuK4KH9pXTv_96d1av_g3zaRK2yf_eu8Hv9liHHXnosG2rXr026glSK6oKv4FQc1KLosygdMdaIKPMaD9DgNUfxWmU2EaKdN0pr8KS8bV7x1--H1D7BN-VJUe</recordid><startdate>200404</startdate><enddate>200404</enddate><creator>Mayer, Melanie L</creator><creator>Pot, Isabelle</creator><creator>Chang, Michael</creator><creator>Xu, Hong</creator><creator>Aneliunas, Victoria</creator><creator>Kwok, Teresa</creator><creator>Newitt, Rick</creator><creator>Aebersold, Ruedi</creator><creator>Boone, Charles</creator><creator>Brown, Grant W</creator><creator>Hieter, Philip</creator><general>The American Society for Cell Biology</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>8FD</scope><scope>FR3</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>200404</creationdate><title>Identification of protein complexes required for efficient sister chromatid cohesion</title><author>Mayer, Melanie L ; 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We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes required for efficient sister chromatid cohesion. We then assessed proficiency of cohesion at three chromosomal loci in strains containing deletions of the genes identified in the ctf8 SGA screen. Deletion of seven genes (CHL1, CSM3, BIM1, KAR3, TOF1, CTF4, and VIK1) resulted in defective sister chromatid cohesion. Mass spectrometric analysis of immunoprecipitated complexes identified a physical association between Kar3p and Vik1p and an interaction between Csm3p and Tof1p that we confirmed by coimmunoprecipitation from cell extracts. These data indicate that synthetic genetic array analysis coupled with specific secondary screens can effectively identify protein complexes functionally related to a reference gene. 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| subjects | Chromatids - ultrastructure Chromosomes, Fungal Epitopes - chemistry Fungal Proteins - physiology Gene Deletion Genetic Techniques Genome, Fungal Green Fluorescent Proteins Luminescent Proteins - metabolism Mass Spectrometry Mutation Oligonucleotide Array Sequence Analysis Precipitin Tests Saccharomyces cerevisiae Saccharomyces cerevisiae - physiology Saccharomyces cerevisiae Proteins - physiology |
| title | Identification of protein complexes required for efficient sister chromatid cohesion |
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