Purified Smc5/6 Complex Exhibits DNA Substrate Recognition and Compaction
Eukaryotic SMC complexes, cohesin, condensin, and Smc5/6, use ATP hydrolysis to power a plethora of functions requiring organization and restructuring of eukaryotic chromosomes in interphase and during mitosis. The Smc5/6 mechanism of action and its activity on DNA are largely unknown. Here we purif...
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| Published in: | Molecular cell Vol. 80; no. 6; p. 1039 |
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| Main Authors: | , , , , , , , , , , , , , |
| Format: | Journal Article |
| Language: | English |
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17.12.2020
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| ISSN: | 1097-4164, 1097-4164 |
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| Abstract | Eukaryotic SMC complexes, cohesin, condensin, and Smc5/6, use ATP hydrolysis to power a plethora of functions requiring organization and restructuring of eukaryotic chromosomes in interphase and during mitosis. The Smc5/6 mechanism of action and its activity on DNA are largely unknown. Here we purified the budding yeast Smc5/6 holocomplex and characterized its core biochemical and biophysical activities. Purified Smc5/6 exhibits DNA-dependent ATP hydrolysis and SUMO E3 ligase activity. We show that Smc5/6 binds DNA topologically with affinity for supercoiled and catenated DNA templates. Employing single-molecule assays to analyze the functional and dynamic characteristics of Smc5/6 bound to DNA, we show that Smc5/6 locks DNA plectonemes and can compact DNA in an ATP-dependent manner. These results demonstrate that the Smc5/6 complex recognizes DNA tertiary structures involving juxtaposed helices and might modulate DNA topology by plectoneme stabilization and local compaction. |
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| AbstractList | Eukaryotic SMC complexes, cohesin, condensin, and Smc5/6, use ATP hydrolysis to power a plethora of functions requiring organization and restructuring of eukaryotic chromosomes in interphase and during mitosis. The Smc5/6 mechanism of action and its activity on DNA are largely unknown. Here we purified the budding yeast Smc5/6 holocomplex and characterized its core biochemical and biophysical activities. Purified Smc5/6 exhibits DNA-dependent ATP hydrolysis and SUMO E3 ligase activity. We show that Smc5/6 binds DNA topologically with affinity for supercoiled and catenated DNA templates. Employing single-molecule assays to analyze the functional and dynamic characteristics of Smc5/6 bound to DNA, we show that Smc5/6 locks DNA plectonemes and can compact DNA in an ATP-dependent manner. These results demonstrate that the Smc5/6 complex recognizes DNA tertiary structures involving juxtaposed helices and might modulate DNA topology by plectoneme stabilization and local compaction.Eukaryotic SMC complexes, cohesin, condensin, and Smc5/6, use ATP hydrolysis to power a plethora of functions requiring organization and restructuring of eukaryotic chromosomes in interphase and during mitosis. The Smc5/6 mechanism of action and its activity on DNA are largely unknown. Here we purified the budding yeast Smc5/6 holocomplex and characterized its core biochemical and biophysical activities. Purified Smc5/6 exhibits DNA-dependent ATP hydrolysis and SUMO E3 ligase activity. We show that Smc5/6 binds DNA topologically with affinity for supercoiled and catenated DNA templates. Employing single-molecule assays to analyze the functional and dynamic characteristics of Smc5/6 bound to DNA, we show that Smc5/6 locks DNA plectonemes and can compact DNA in an ATP-dependent manner. These results demonstrate that the Smc5/6 complex recognizes DNA tertiary structures involving juxtaposed helices and might modulate DNA topology by plectoneme stabilization and local compaction. Eukaryotic SMC complexes, cohesin, condensin, and Smc5/6, use ATP hydrolysis to power a plethora of functions requiring organization and restructuring of eukaryotic chromosomes in interphase and during mitosis. The Smc5/6 mechanism of action and its activity on DNA are largely unknown. Here we purified the budding yeast Smc5/6 holocomplex and characterized its core biochemical and biophysical activities. Purified Smc5/6 exhibits DNA-dependent ATP hydrolysis and SUMO E3 ligase activity. We show that Smc5/6 binds DNA topologically with affinity for supercoiled and catenated DNA templates. Employing single-molecule assays to analyze the functional and dynamic characteristics of Smc5/6 bound to DNA, we show that Smc5/6 locks DNA plectonemes and can compact DNA in an ATP-dependent manner. These results demonstrate that the Smc5/6 complex recognizes DNA tertiary structures involving juxtaposed helices and might modulate DNA topology by plectoneme stabilization and local compaction. |
| Author | Gutierrez-Escribano, Pilar Rappsilber, Juri Aragon, Luis Montoya, Alex Solé-Soler, Roger Morris, Kyle O'Reilly, Francis J Hormeño, Silvia Madariaga-Marcos, Julene Aicart-Ramos, Clara Aramayo, Ricardo Torres-Rosell, Jordi Kramer, Holger Moreno-Herrero, Fernando |
| Author_xml | – sequence: 1 givenname: Pilar surname: Gutierrez-Escribano fullname: Gutierrez-Escribano, Pilar organization: Cell Cycle Group, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK – sequence: 2 givenname: Silvia surname: Hormeño fullname: Hormeño, Silvia organization: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Madrid, Spain – sequence: 3 givenname: Julene surname: Madariaga-Marcos fullname: Madariaga-Marcos, Julene organization: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Madrid, Spain – sequence: 4 givenname: Roger surname: Solé-Soler fullname: Solé-Soler, Roger organization: Institut de Recerca Biomèdica de Lleida (IRBLLEIDA), Department of Ciències Mèdiques Bàsiques, Universitat de Lleida, Lleida, Spain – sequence: 5 givenname: Francis J surname: O'Reilly fullname: O'Reilly, Francis J organization: Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, 13355 Berlin, Germany; Wellcome Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, UK – sequence: 6 givenname: Kyle surname: Morris fullname: Morris, Kyle organization: Microscopy Facility, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK – sequence: 7 givenname: Clara surname: Aicart-Ramos fullname: Aicart-Ramos, Clara organization: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Madrid, Spain – sequence: 8 givenname: Ricardo surname: Aramayo fullname: Aramayo, Ricardo organization: Microscopy Facility, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK – sequence: 9 givenname: Alex surname: Montoya fullname: Montoya, Alex organization: Biological Mass Spectrometry and Proteomics Facility, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK – sequence: 10 givenname: Holger surname: Kramer fullname: Kramer, Holger organization: Biological Mass Spectrometry and Proteomics Facility, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK – sequence: 11 givenname: Juri surname: Rappsilber fullname: Rappsilber, Juri organization: Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, 13355 Berlin, Germany; Wellcome Centre for Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, UK – sequence: 12 givenname: Jordi surname: Torres-Rosell fullname: Torres-Rosell, Jordi organization: Institut de Recerca Biomèdica de Lleida (IRBLLEIDA), Department of Ciències Mèdiques Bàsiques, Universitat de Lleida, Lleida, Spain – sequence: 13 givenname: Fernando surname: Moreno-Herrero fullname: Moreno-Herrero, Fernando email: fernando.moreno@cnb.csic.es organization: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Madrid, Spain. Electronic address: fernando.moreno@cnb.csic.es – sequence: 14 givenname: Luis surname: Aragon fullname: Aragon, Luis email: luis.aragon@lms.mrc.ac.uk organization: Cell Cycle Group, MRC London Institute of Medical Sciences (LMS), Du Cane Road, London W12 0NN, UK. Electronic address: luis.aragon@lms.mrc.ac.uk |
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| Keywords | electron mmicroscopy single-molecule magnetic tweezers Smc5/6 holocomplex plectoneme stabilisation Smc5/6 purification DNA compaction DNA substrate recognition |
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| SubjectTerms | Adenosine Triphosphatases - genetics Biophysical Phenomena Cell Cycle Proteins - genetics Cell Cycle Proteins - ultrastructure Chromosomal Proteins, Non-Histone - genetics Chromosomal Proteins, Non-Histone - ultrastructure Cohesins DNA-Binding Proteins - genetics Humans Interphase - genetics Mitosis - genetics Multiprotein Complexes - genetics Multiprotein Complexes - ultrastructure Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - ultrastructure Saccharomyces cerevisiae Proteins - genetics Saccharomyces cerevisiae Proteins - ultrastructure Sumoylation - genetics |
| Title | Purified Smc5/6 Complex Exhibits DNA Substrate Recognition and Compaction |
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