Interaction of DNA Fragmentation Factor (DFF) with DNA Reveals an Unprecedented Mechanism for Nuclease Inhibition and Suggests That DFF Can Be Activated in a DNA-bound State
DNA fragmentation factor (DFF) is a complex of the DNase DFF40 (CAD) and its chaperone/inhibitor DFF45 (ICAD-L) that can be activated during apoptosis to induce DNA fragmentation. Here, we demonstrate that DFF directly binds to DNA in vitro without promoting DNA cleavage. DNA binding by DFF is media...
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| Veröffentlicht in: | The Journal of biological chemistry 2005-02, Vol.280 (7), p.6005-6015 |
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| container_title | The Journal of biological chemistry |
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| creator | Korn, Christian Scholz, Sebastian R. Gimadutdinow, Oleg Lurz, Rudi Pingoud, Alfred Meiss, Gregor |
| description | DNA fragmentation factor (DFF) is a complex of the DNase DFF40 (CAD) and its chaperone/inhibitor DFF45 (ICAD-L) that can be activated during apoptosis to induce DNA fragmentation. Here, we demonstrate that DFF directly binds to DNA in vitro without promoting DNA cleavage. DNA binding by DFF is mediated by the nuclease subunit, which can also form stable DNA complexes after release from DFF. Recombinant and reconstituted DFF is catalytically inactive yet proficient in DNA binding, demonstrating that the nuclease subunit in DFF is inhibited in DNA cleavage but not in DNA binding, revealing an unprecedented mode of nuclease inhibition. Activation of DFF in the presence of naked DNA or isolated nuclei stimulates DNA degradation by released DFF40 (CAD). In transfected HeLa cells transiently expressed DFF associates with chromatin, suggesting that DFF could be activated during apoptosis in a DNA-bound state. |
| doi_str_mv | 10.1074/jbc.M413035200 |
| format | Article |
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Here, we demonstrate that DFF directly binds to DNA in vitro without promoting DNA cleavage. DNA binding by DFF is mediated by the nuclease subunit, which can also form stable DNA complexes after release from DFF. Recombinant and reconstituted DFF is catalytically inactive yet proficient in DNA binding, demonstrating that the nuclease subunit in DFF is inhibited in DNA cleavage but not in DNA binding, revealing an unprecedented mode of nuclease inhibition. Activation of DFF in the presence of naked DNA or isolated nuclei stimulates DNA degradation by released DFF40 (CAD). 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In transfected HeLa cells transiently expressed DFF associates with chromatin, suggesting that DFF could be activated during apoptosis in a DNA-bound state.</description><subject>Animals</subject><subject>Apoptosis</subject><subject>Apoptosis Regulatory Proteins</subject><subject>Base Sequence</subject><subject>Caspase 3</subject><subject>Caspases - metabolism</subject><subject>Chromatin - metabolism</subject><subject>Deoxyribonucleases - antagonists & inhibitors</subject><subject>Deoxyribonucleases - chemistry</subject><subject>Deoxyribonucleases - genetics</subject><subject>Deoxyribonucleases - metabolism</subject><subject>DNA - genetics</subject><subject>DNA - metabolism</subject><subject>DNA - ultrastructure</subject><subject>Enzyme Activation</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>Mice</subject><subject>Microscopy, Electron, Transmission</subject><subject>Models, Biological</subject><subject>Models, Molecular</subject><subject>Poly-ADP-Ribose Binding Proteins</subject><subject>Protein Binding</subject><subject>Protein Conformation</subject><subject>Protein Subunits - antagonists & inhibitors</subject><subject>Protein Subunits - chemistry</subject><subject>Protein Subunits - genetics</subject><subject>Protein Subunits - metabolism</subject><subject>Protein Transport</subject><subject>Proteins - genetics</subject><subject>Proteins - metabolism</subject><subject>Recombinant Proteins - chemistry</subject><subject>Recombinant Proteins - genetics</subject><subject>Recombinant Proteins - metabolism</subject><subject>Substrate Specificity</subject><subject>Transfection</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>eNqFkUtv1DAUhS0EokNhyxJZLBAsMtjOy14O0wZGaosErcTOcpybiauJM9jJVPyo_kfuPKSuEN5Ysr9z7tE9hLzlbM5ZmX2-r-38OuMpS3PB2DMy40ymSZrzX8_JjDHBEyVyeUZexXjP8GSKvyRnPM9LgdCMPK78CMHY0Q2eDi29uFnQKph1D340h8cKP4dAP15U1Sf64MbuwPyAHZhNpMbTO78NYKFBBTT0GmxnvIs9bVF1M9kNmAh05TtXu4Oh8Q39Oa3XEMdIbzszUrSmS3T6AnSBSXZmb-SQ3I9K6mHaKzAOvCYvWpwKb073ObmrLm-X35Kr719Xy8VVYrOMjUkhm1ylPFUFr9OiZkrWUKuWGytMkUqjeNkCCFlAVigp80LVyrSyAcWEkLjLc_Lh6LsNw-8Jg-reRQubjfEwTFEXZSbSkqX_BXkpsZuiRHB-BG0YYgzQ6m1wvQl_NGd636TGJvVTkyh4d3Ke6h6aJ_xUHQLvj0Dn1t2DC6BrN9gOei0k06UuGMsRkkcIcFs7B0FH68BjWyiwo24G968AfwFarbZA</recordid><startdate>20050218</startdate><enddate>20050218</enddate><creator>Korn, Christian</creator><creator>Scholz, Sebastian R.</creator><creator>Gimadutdinow, Oleg</creator><creator>Lurz, Rudi</creator><creator>Pingoud, Alfred</creator><creator>Meiss, Gregor</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><scope>7X8</scope></search><sort><creationdate>20050218</creationdate><title>Interaction of DNA Fragmentation Factor (DFF) with DNA Reveals an Unprecedented Mechanism for Nuclease Inhibition and Suggests That DFF Can Be Activated in a DNA-bound State</title><author>Korn, Christian ; 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Here, we demonstrate that DFF directly binds to DNA in vitro without promoting DNA cleavage. DNA binding by DFF is mediated by the nuclease subunit, which can also form stable DNA complexes after release from DFF. Recombinant and reconstituted DFF is catalytically inactive yet proficient in DNA binding, demonstrating that the nuclease subunit in DFF is inhibited in DNA cleavage but not in DNA binding, revealing an unprecedented mode of nuclease inhibition. Activation of DFF in the presence of naked DNA or isolated nuclei stimulates DNA degradation by released DFF40 (CAD). In transfected HeLa cells transiently expressed DFF associates with chromatin, suggesting that DFF could be activated during apoptosis in a DNA-bound state.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>15572351</pmid><doi>10.1074/jbc.M413035200</doi><tpages>11</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Animals Apoptosis Apoptosis Regulatory Proteins Base Sequence Caspase 3 Caspases - metabolism Chromatin - metabolism Deoxyribonucleases - antagonists & inhibitors Deoxyribonucleases - chemistry Deoxyribonucleases - genetics Deoxyribonucleases - metabolism DNA - genetics DNA - metabolism DNA - ultrastructure Enzyme Activation HeLa Cells Humans Mice Microscopy, Electron, Transmission Models, Biological Models, Molecular Poly-ADP-Ribose Binding Proteins Protein Binding Protein Conformation Protein Subunits - antagonists & inhibitors Protein Subunits - chemistry Protein Subunits - genetics Protein Subunits - metabolism Protein Transport Proteins - genetics Proteins - metabolism Recombinant Proteins - chemistry Recombinant Proteins - genetics Recombinant Proteins - metabolism Substrate Specificity Transfection |
| title | Interaction of DNA Fragmentation Factor (DFF) with DNA Reveals an Unprecedented Mechanism for Nuclease Inhibition and Suggests That DFF Can Be Activated in a DNA-bound State |
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