CDK-dependent complex formation between replication proteins Dpb11, Sld2, Pol (epsilon}, and GINS in budding yeast
Eukaryotic chromosomal DNA replication requires cyclin-dependent kinase (CDK) activity. CDK phosphorylates two yeast replication proteins, Sld3 and Sld2, both of which bind to Dpb11 when phosphorylated. These phosphorylation-dependent interactions are essential and are the minimal requirements for C...
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Veröffentlicht in: | Genes & development 2010-03, Vol.24 (6), p.602-612 |
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creator | Muramatsu, Sachiko Hirai, Kazuyuki Tak, Yon-Soo Kamimura, Yoichiro Araki, Hiroyuki |
description | Eukaryotic chromosomal DNA replication requires cyclin-dependent kinase (CDK) activity. CDK phosphorylates two yeast replication proteins, Sld3 and Sld2, both of which bind to Dpb11 when phosphorylated. These phosphorylation-dependent interactions are essential and are the minimal requirements for CDK-dependent activation of DNA replication. However, how these interactions activate DNA replication has not been elucidated. Here, we show that CDK promotes the formation of a newly identified fragile complex, the preloading complex (pre-LC) containing DNA polymerase epsilon (Pol epsilon), GINS, Sld2, and Dpb11. Formation of the pre-LC requires phosphorylation of Sld2 by CDK, but is independent of DNA replication, protein association with replication origins, and Dbf4-dependent Cdc7 kinase, which is also essential for the activation of DNA replication. We also demonstrate that Pol epsilon, GINS, Dpb11, and CDK-phosphorylated Sld2 form a complex in vitro. The genetic interactions between Pol epsilon, GINS, Sld2, and Dpb11 suggest further that they form an essential complex in cells. We propose that CDK regulates the initiation of DNA replication in budding yeast through formation of the pre-LC. |
doi_str_mv | 10.1101/gad.1883410 |
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CDK phosphorylates two yeast replication proteins, Sld3 and Sld2, both of which bind to Dpb11 when phosphorylated. These phosphorylation-dependent interactions are essential and are the minimal requirements for CDK-dependent activation of DNA replication. However, how these interactions activate DNA replication has not been elucidated. Here, we show that CDK promotes the formation of a newly identified fragile complex, the preloading complex (pre-LC) containing DNA polymerase epsilon (Pol epsilon), GINS, Sld2, and Dpb11. Formation of the pre-LC requires phosphorylation of Sld2 by CDK, but is independent of DNA replication, protein association with replication origins, and Dbf4-dependent Cdc7 kinase, which is also essential for the activation of DNA replication. We also demonstrate that Pol epsilon, GINS, Dpb11, and CDK-phosphorylated Sld2 form a complex in vitro. The genetic interactions between Pol epsilon, GINS, Sld2, and Dpb11 suggest further that they form an essential complex in cells. 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CDK phosphorylates two yeast replication proteins, Sld3 and Sld2, both of which bind to Dpb11 when phosphorylated. These phosphorylation-dependent interactions are essential and are the minimal requirements for CDK-dependent activation of DNA replication. However, how these interactions activate DNA replication has not been elucidated. Here, we show that CDK promotes the formation of a newly identified fragile complex, the preloading complex (pre-LC) containing DNA polymerase epsilon (Pol epsilon), GINS, Sld2, and Dpb11. Formation of the pre-LC requires phosphorylation of Sld2 by CDK, but is independent of DNA replication, protein association with replication origins, and Dbf4-dependent Cdc7 kinase, which is also essential for the activation of DNA replication. We also demonstrate that Pol epsilon, GINS, Dpb11, and CDK-phosphorylated Sld2 form a complex in vitro. The genetic interactions between Pol epsilon, GINS, Sld2, and Dpb11 suggest further that they form an essential complex in cells. We propose that CDK regulates the initiation of DNA replication in budding yeast through formation of the pre-LC.</description><subject>Cell Cycle</subject><subject>Cell Cycle Proteins - metabolism</subject><subject>Cyclin-Dependent Kinases - metabolism</subject><subject>DNA Polymerase II - metabolism</subject><subject>Multiprotein Complexes - metabolism</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>Saccharomyces cerevisiae - growth & development</subject><subject>Saccharomyces cerevisiae - metabolism</subject><subject>Saccharomyces cerevisiae Proteins - metabolism</subject><issn>1549-5477</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2010</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo1kMFLwzAYxYMgbk5P3iU3PawzX9M2zVE2ncOhwnYvSfN1VNI0Ni26g_-7g83Tg8ePH49HyA2wGQCDh50yM8hzngA7I2NIExmliRAjchnCJ2MsY1l2QUYxizlwEGPSzRevkUGPzqDradk23uIPrdquUX3dOqqx_0Z0tENv6_LY-a7tsXaBLrwGmNKNNfGUfrSW3qMPtW3d75QqZ-hy9bah9UEyGFO7Hd2jCv0VOa-UDXh9ygnZPj9t5y_R-n25mj-uIx8D6yMuVcxKI4VWINMKNcYJxJmCSuqUG5mXGiRHmYpMlDnnPNVaVVDmOUMBmk_I3VF7WPs1YOiLpg4lWqsctkMoBOcCEi7SA3l7IgfdoCl8Vzeq2xf_L_E_mDBmfg</recordid><startdate>20100315</startdate><enddate>20100315</enddate><creator>Muramatsu, Sachiko</creator><creator>Hirai, Kazuyuki</creator><creator>Tak, Yon-Soo</creator><creator>Kamimura, Yoichiro</creator><creator>Araki, Hiroyuki</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>7X8</scope></search><sort><creationdate>20100315</creationdate><title>CDK-dependent complex formation between replication proteins Dpb11, Sld2, Pol (epsilon}, and GINS in budding yeast</title><author>Muramatsu, Sachiko ; Hirai, Kazuyuki ; Tak, Yon-Soo ; Kamimura, Yoichiro ; Araki, Hiroyuki</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-p210t-39a20cd97ba195febe24126a1f9b53d98cb193e95767c83335bbaf1c880e71b3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2010</creationdate><topic>Cell Cycle</topic><topic>Cell Cycle Proteins - metabolism</topic><topic>Cyclin-Dependent Kinases - metabolism</topic><topic>DNA Polymerase II - metabolism</topic><topic>Multiprotein Complexes - metabolism</topic><topic>Saccharomyces cerevisiae - genetics</topic><topic>Saccharomyces cerevisiae - growth & development</topic><topic>Saccharomyces cerevisiae - metabolism</topic><topic>Saccharomyces cerevisiae Proteins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Muramatsu, Sachiko</creatorcontrib><creatorcontrib>Hirai, Kazuyuki</creatorcontrib><creatorcontrib>Tak, Yon-Soo</creatorcontrib><creatorcontrib>Kamimura, Yoichiro</creatorcontrib><creatorcontrib>Araki, Hiroyuki</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>MEDLINE - Academic</collection><jtitle>Genes & development</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Muramatsu, Sachiko</au><au>Hirai, Kazuyuki</au><au>Tak, Yon-Soo</au><au>Kamimura, Yoichiro</au><au>Araki, Hiroyuki</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>CDK-dependent complex formation between replication proteins Dpb11, Sld2, Pol (epsilon}, and GINS in budding yeast</atitle><jtitle>Genes & development</jtitle><addtitle>Genes Dev</addtitle><date>2010-03-15</date><risdate>2010</risdate><volume>24</volume><issue>6</issue><spage>602</spage><epage>612</epage><pages>602-612</pages><eissn>1549-5477</eissn><abstract>Eukaryotic chromosomal DNA replication requires cyclin-dependent kinase (CDK) activity. CDK phosphorylates two yeast replication proteins, Sld3 and Sld2, both of which bind to Dpb11 when phosphorylated. These phosphorylation-dependent interactions are essential and are the minimal requirements for CDK-dependent activation of DNA replication. However, how these interactions activate DNA replication has not been elucidated. Here, we show that CDK promotes the formation of a newly identified fragile complex, the preloading complex (pre-LC) containing DNA polymerase epsilon (Pol epsilon), GINS, Sld2, and Dpb11. Formation of the pre-LC requires phosphorylation of Sld2 by CDK, but is independent of DNA replication, protein association with replication origins, and Dbf4-dependent Cdc7 kinase, which is also essential for the activation of DNA replication. We also demonstrate that Pol epsilon, GINS, Dpb11, and CDK-phosphorylated Sld2 form a complex in vitro. 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subjects | Cell Cycle Cell Cycle Proteins - metabolism Cyclin-Dependent Kinases - metabolism DNA Polymerase II - metabolism Multiprotein Complexes - metabolism Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - growth & development Saccharomyces cerevisiae - metabolism Saccharomyces cerevisiae Proteins - metabolism |
title | CDK-dependent complex formation between replication proteins Dpb11, Sld2, Pol (epsilon}, and GINS in budding yeast |
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