Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks
DNA-dependent protein kinase (DNA-PK), consisting of Ku and DNA-PKcs subunits, is the key component of the non-homologous end-joining (NHEJ) pathway of DNA double strand break (DSB) repair. Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclu...
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| Veröffentlicht in: | The Journal of biological chemistry 2005-04, Vol.280 (15), p.14709-14715 |
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| container_title | The Journal of biological chemistry |
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| creator | Chen, Benjamin P.C. Chan, Doug W. Kobayashi, Junya Burma, Sandeep Asaithamby, Aroumougame Morotomi-Yano, Keiko Botvinick, Elliot Qin, Jun Chen, David J. |
| description | DNA-dependent protein kinase (DNA-PK), consisting of Ku and DNA-PKcs subunits, is the key component of the non-homologous end-joining (NHEJ) pathway of DNA double strand break (DSB) repair. Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclusively identified except for an autophosphorylation site on DNA-PKcs itself (threonine 2609). Here we report the ionizing radiation (IR)-induced autophosphorylation of DNA-PKcs at a novel site, serine 2056, the phosphorylation of which is required for the repair of DSBs by NHEJ. Interestingly, IR-induced DNA-PKcs autophosphorylation is regulated in a cell cycle-dependent manner with attenuated phosphorylation in the S phase. In contrast, DNA replication-associated DSBs resulted in DNA-PKcs autophosphorylation and localization to DNA damage sites. These results indicate that although IR-induced DNA-PKcs phosphorylation is attenuated in the S phase, DNA-PKcs is preferentially activated by the physiologically relevant DNA replication-associated DSBs at the sites of DNA synthesis. |
| doi_str_mv | 10.1074/jbc.M408827200 |
| format | Article |
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Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclusively identified except for an autophosphorylation site on DNA-PKcs itself (threonine 2609). Here we report the ionizing radiation (IR)-induced autophosphorylation of DNA-PKcs at a novel site, serine 2056, the phosphorylation of which is required for the repair of DSBs by NHEJ. Interestingly, IR-induced DNA-PKcs autophosphorylation is regulated in a cell cycle-dependent manner with attenuated phosphorylation in the S phase. In contrast, DNA replication-associated DSBs resulted in DNA-PKcs autophosphorylation and localization to DNA damage sites. These results indicate that although IR-induced DNA-PKcs phosphorylation is attenuated in the S phase, DNA-PKcs is preferentially activated by the physiologically relevant DNA replication-associated DSBs at the sites of DNA synthesis.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M408827200</identifier><identifier>PMID: 15677476</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Amino Acid Sequence ; Cell Cycle ; Cell Line ; Cell Line, Tumor ; Cell Nucleus - metabolism ; DNA Damage ; DNA Repair ; DNA-Activated Protein Kinase ; DNA-Binding Proteins - chemistry ; DNA-Binding Proteins - metabolism ; Electrophoresis, Polyacrylamide Gel ; Genetic Vectors ; HeLa Cells ; Humans ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; Nuclear Proteins ; Phosphorylation ; Protein-Serine-Threonine Kinases - chemistry ; Protein-Serine-Threonine Kinases - metabolism ; Radiation, Ionizing ; S Phase ; Serine - chemistry ; Threonine - chemistry</subject><ispartof>The Journal of biological chemistry, 2005-04, Vol.280 (15), p.14709-14715</ispartof><rights>2005 © 2005 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c442t-2635f752b9588d7fa64b46bb4718b5ed23b8101e710307fad588b3f3a500dc883</citedby><cites>FETCH-LOGICAL-c442t-2635f752b9588d7fa64b46bb4718b5ed23b8101e710307fad588b3f3a500dc883</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15677476$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Chen, Benjamin P.C.</creatorcontrib><creatorcontrib>Chan, Doug W.</creatorcontrib><creatorcontrib>Kobayashi, Junya</creatorcontrib><creatorcontrib>Burma, Sandeep</creatorcontrib><creatorcontrib>Asaithamby, Aroumougame</creatorcontrib><creatorcontrib>Morotomi-Yano, Keiko</creatorcontrib><creatorcontrib>Botvinick, Elliot</creatorcontrib><creatorcontrib>Qin, Jun</creatorcontrib><creatorcontrib>Chen, David J.</creatorcontrib><title>Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>DNA-dependent protein kinase (DNA-PK), consisting of Ku and DNA-PKcs subunits, is the key component of the non-homologous end-joining (NHEJ) pathway of DNA double strand break (DSB) repair. Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclusively identified except for an autophosphorylation site on DNA-PKcs itself (threonine 2609). Here we report the ionizing radiation (IR)-induced autophosphorylation of DNA-PKcs at a novel site, serine 2056, the phosphorylation of which is required for the repair of DSBs by NHEJ. Interestingly, IR-induced DNA-PKcs autophosphorylation is regulated in a cell cycle-dependent manner with attenuated phosphorylation in the S phase. In contrast, DNA replication-associated DSBs resulted in DNA-PKcs autophosphorylation and localization to DNA damage sites. These results indicate that although IR-induced DNA-PKcs phosphorylation is attenuated in the S phase, DNA-PKcs is preferentially activated by the physiologically relevant DNA replication-associated DSBs at the sites of DNA synthesis.</description><subject>Amino Acid Sequence</subject><subject>Cell Cycle</subject><subject>Cell Line</subject><subject>Cell Line, Tumor</subject><subject>Cell Nucleus - metabolism</subject><subject>DNA Damage</subject><subject>DNA Repair</subject><subject>DNA-Activated Protein Kinase</subject><subject>DNA-Binding Proteins - chemistry</subject><subject>DNA-Binding Proteins - metabolism</subject><subject>Electrophoresis, Polyacrylamide Gel</subject><subject>Genetic Vectors</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>Molecular Sequence Data</subject><subject>Mutagenesis, Site-Directed</subject><subject>Nuclear Proteins</subject><subject>Phosphorylation</subject><subject>Protein-Serine-Threonine Kinases - chemistry</subject><subject>Protein-Serine-Threonine Kinases - metabolism</subject><subject>Radiation, Ionizing</subject><subject>S Phase</subject><subject>Serine - chemistry</subject><subject>Threonine - chemistry</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kEtv1DAURi0EokNhyxJ5gdhlsB07dpZlpoWqBSoeEjvLjxviksTBzoDm3-PRjNQV3ljyd-6n64PQS0rWlEj-9t669UdOlGKSEfIIrShRdVUL-uMxWhHCaNUyoc7Qs5zvSTm8pU_RGRWNlFw2KzRuYBjwZu8GwFuYYfIwOcCxw9tPF5U_vSz4LsUFwoRvwmQy4Ls-5rmPaT-YJcQJl-QL5DlOJVviYRZv486W0q9LMpPH7xKYX_k5etKZIcOL032Ovl9dftt8qG4_v7_eXNxWjnO2VKypRScFs61QysvONNzyxlouqbICPKutooSCpKQmJfYFs3VXG0GId0rV5-jNsXdO8fcO8qLHkF35qZkg7rKmkpfqlhVwfQRdijkn6PScwmjSXlOiD4J1EawfBJeBV6fmnR3BP-AnowV4fQT68LP_GxJoG6LrYdRMlUqhKZekLZg6YlA0_AmQdHbhoN6XEbdoH8P_VvgHaYKUdw</recordid><startdate>20050415</startdate><enddate>20050415</enddate><creator>Chen, Benjamin P.C.</creator><creator>Chan, Doug W.</creator><creator>Kobayashi, Junya</creator><creator>Burma, Sandeep</creator><creator>Asaithamby, Aroumougame</creator><creator>Morotomi-Yano, Keiko</creator><creator>Botvinick, Elliot</creator><creator>Qin, Jun</creator><creator>Chen, David J.</creator><general>Elsevier Inc</general><general>American Society for Biochemistry and Molecular Biology</general><scope>6I.</scope><scope>AAFTH</scope><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>7TM</scope></search><sort><creationdate>20050415</creationdate><title>Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks</title><author>Chen, Benjamin P.C. ; Chan, Doug W. ; Kobayashi, Junya ; Burma, Sandeep ; Asaithamby, Aroumougame ; Morotomi-Yano, Keiko ; Botvinick, Elliot ; Qin, Jun ; Chen, David J.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c442t-2635f752b9588d7fa64b46bb4718b5ed23b8101e710307fad588b3f3a500dc883</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2005</creationdate><topic>Amino Acid Sequence</topic><topic>Cell Cycle</topic><topic>Cell Line</topic><topic>Cell Line, Tumor</topic><topic>Cell Nucleus - metabolism</topic><topic>DNA Damage</topic><topic>DNA Repair</topic><topic>DNA-Activated Protein Kinase</topic><topic>DNA-Binding Proteins - chemistry</topic><topic>DNA-Binding Proteins - metabolism</topic><topic>Electrophoresis, Polyacrylamide Gel</topic><topic>Genetic Vectors</topic><topic>HeLa Cells</topic><topic>Humans</topic><topic>Molecular Sequence Data</topic><topic>Mutagenesis, Site-Directed</topic><topic>Nuclear Proteins</topic><topic>Phosphorylation</topic><topic>Protein-Serine-Threonine Kinases - chemistry</topic><topic>Protein-Serine-Threonine Kinases - metabolism</topic><topic>Radiation, Ionizing</topic><topic>S Phase</topic><topic>Serine - chemistry</topic><topic>Threonine - chemistry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Chen, Benjamin P.C.</creatorcontrib><creatorcontrib>Chan, Doug W.</creatorcontrib><creatorcontrib>Kobayashi, Junya</creatorcontrib><creatorcontrib>Burma, Sandeep</creatorcontrib><creatorcontrib>Asaithamby, Aroumougame</creatorcontrib><creatorcontrib>Morotomi-Yano, Keiko</creatorcontrib><creatorcontrib>Botvinick, Elliot</creatorcontrib><creatorcontrib>Qin, Jun</creatorcontrib><creatorcontrib>Chen, David J.</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Chen, Benjamin P.C.</au><au>Chan, Doug W.</au><au>Kobayashi, Junya</au><au>Burma, Sandeep</au><au>Asaithamby, Aroumougame</au><au>Morotomi-Yano, Keiko</au><au>Botvinick, Elliot</au><au>Qin, Jun</au><au>Chen, David J.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>2005-04-15</date><risdate>2005</risdate><volume>280</volume><issue>15</issue><spage>14709</spage><epage>14715</epage><pages>14709-14715</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>DNA-dependent protein kinase (DNA-PK), consisting of Ku and DNA-PKcs subunits, is the key component of the non-homologous end-joining (NHEJ) pathway of DNA double strand break (DSB) repair. Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclusively identified except for an autophosphorylation site on DNA-PKcs itself (threonine 2609). Here we report the ionizing radiation (IR)-induced autophosphorylation of DNA-PKcs at a novel site, serine 2056, the phosphorylation of which is required for the repair of DSBs by NHEJ. Interestingly, IR-induced DNA-PKcs autophosphorylation is regulated in a cell cycle-dependent manner with attenuated phosphorylation in the S phase. In contrast, DNA replication-associated DSBs resulted in DNA-PKcs autophosphorylation and localization to DNA damage sites. 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| subjects | Amino Acid Sequence Cell Cycle Cell Line Cell Line, Tumor Cell Nucleus - metabolism DNA Damage DNA Repair DNA-Activated Protein Kinase DNA-Binding Proteins - chemistry DNA-Binding Proteins - metabolism Electrophoresis, Polyacrylamide Gel Genetic Vectors HeLa Cells Humans Molecular Sequence Data Mutagenesis, Site-Directed Nuclear Proteins Phosphorylation Protein-Serine-Threonine Kinases - chemistry Protein-Serine-Threonine Kinases - metabolism Radiation, Ionizing S Phase Serine - chemistry Threonine - chemistry |
| title | Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks |
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