Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ
DNA synthesis across lesions during genomic replication requires concerted actions of specialized DNA polymerases in a potentially mutagenic process known as translesion synthesis. Current models suggest that translesion synthesis in mammalian cells is achieved in two sequential steps, with a Y-fami...
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| Format: | Article |
| Language: | en_US |
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American Society for Biochemistry and Molecular Biology (ASBMB)
2014
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| Online Access: | http://hdl.handle.net/1721.1/86395 https://orcid.org/0000-0001-7243-8261 |
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| author | Wojtaszek, Jessica Lee, Chul-Jin D'Souza, Sanjay Victor Minesinger, Brenda Kim, Hyungjin D'Andrea, Alan D. Walker, Graham C. Zhou, Pei |
| author2 | Massachusetts Institute of Technology. Department of Biology |
| author_facet | Massachusetts Institute of Technology. Department of Biology Wojtaszek, Jessica Lee, Chul-Jin D'Souza, Sanjay Victor Minesinger, Brenda Kim, Hyungjin D'Andrea, Alan D. Walker, Graham C. Zhou, Pei |
| author_sort | Wojtaszek, Jessica |
| collection | MIT |
| description | DNA synthesis across lesions during genomic replication requires concerted actions of specialized DNA polymerases in a potentially mutagenic process known as translesion synthesis. Current models suggest that translesion synthesis in mammalian cells is achieved in two sequential steps, with a Y-family DNA polymerase (κ, η, ι, or Rev1) inserting a nucleotide opposite the lesion and with the heterodimeric B-family polymerase ζ, consisting of the catalytic Rev3 subunit and the accessory Rev7 subunit, replacing the insertion polymerase to carry out primer extension past the lesion. Effective translesion synthesis in vertebrates requires the scaffolding function of the C-terminal domain (CTD) of Rev1 that interacts with the Rev1-interacting region of polymerases κ, η, and ι and with the Rev7 subunit of polymerase ζ. We report the purification and structure determination of a quaternary translesion polymerase complex consisting of the Rev1 CTD, the heterodimeric Pol ζ complex, and the Pol κ Rev1-interacting region. Yeast two-hybrid assays were employed to identify important interface residues of the translesion polymerase complex. The structural elucidation of such a quaternary translesion polymerase complex encompassing both insertion and extension polymerases bridged by the Rev1 CTD provides the first molecular explanation of the essential scaffolding function of Rev1 and highlights the Rev1 CTD as a promising target for developing novel cancer therapeutics to suppress translesion synthesis. Our studies support the notion that vertebrate insertion and extension polymerases could structurally cooperate within a megatranslesion polymerase complex (translesionsome) nucleated by Rev1 to achieve efficient lesion bypass without incurring an additional switching mechanism. |
| first_indexed | 2024-09-23T08:43:28Z |
| format | Article |
| id | mit-1721.1/86395 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T08:43:28Z |
| publishDate | 2014 |
| publisher | American Society for Biochemistry and Molecular Biology (ASBMB) |
| record_format | dspace |
| spelling | mit-1721.1/863952022-09-23T14:06:39Z Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ Wojtaszek, Jessica Lee, Chul-Jin D'Souza, Sanjay Victor Minesinger, Brenda Kim, Hyungjin D'Andrea, Alan D. Walker, Graham C. Zhou, Pei Massachusetts Institute of Technology. Department of Biology Walker, Graham C. D'Souza, Sanjay Victor Minesinger, Brenda Walker, Graham C. DNA synthesis across lesions during genomic replication requires concerted actions of specialized DNA polymerases in a potentially mutagenic process known as translesion synthesis. Current models suggest that translesion synthesis in mammalian cells is achieved in two sequential steps, with a Y-family DNA polymerase (κ, η, ι, or Rev1) inserting a nucleotide opposite the lesion and with the heterodimeric B-family polymerase ζ, consisting of the catalytic Rev3 subunit and the accessory Rev7 subunit, replacing the insertion polymerase to carry out primer extension past the lesion. Effective translesion synthesis in vertebrates requires the scaffolding function of the C-terminal domain (CTD) of Rev1 that interacts with the Rev1-interacting region of polymerases κ, η, and ι and with the Rev7 subunit of polymerase ζ. We report the purification and structure determination of a quaternary translesion polymerase complex consisting of the Rev1 CTD, the heterodimeric Pol ζ complex, and the Pol κ Rev1-interacting region. Yeast two-hybrid assays were employed to identify important interface residues of the translesion polymerase complex. The structural elucidation of such a quaternary translesion polymerase complex encompassing both insertion and extension polymerases bridged by the Rev1 CTD provides the first molecular explanation of the essential scaffolding function of Rev1 and highlights the Rev1 CTD as a promising target for developing novel cancer therapeutics to suppress translesion synthesis. Our studies support the notion that vertebrate insertion and extension polymerases could structurally cooperate within a megatranslesion polymerase complex (translesionsome) nucleated by Rev1 to achieve efficient lesion bypass without incurring an additional switching mechanism. National Institute of General Medical Sciences (U.S.) (NIH) Grant GM-079376) National Institute of Environmental Health Sciences (NIH GrantES-015818) National Institutes of Health (U.S.) (grant R01DK43889) National Institutes of Health (U.S.) (grant R37HL52725) National Institutes of Health (U.S.) (grant RC4DK090913) Alexander and Margaret Stewart Trust American Cancer Society (Research professorship) 2014-05-02T19:57:03Z 2014-05-02T19:57:03Z 2012-08 2012-07 Article http://purl.org/eprint/type/JournalArticle 0021-9258 1083-351X http://hdl.handle.net/1721.1/86395 Wojtaszek, J., C.-J. Lee, S. D’Souza, B. Minesinger, H. Kim, A. D. D’Andrea, G. C. Walker, and P. Zhou. “Structural Basis of Rev1-Mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ.” Journal of Biological Chemistry 287, no. 40 (September 28, 2012): 33836–33846. https://orcid.org/0000-0001-7243-8261 en_US http://dx.doi.org/10.1074/jbc.M112.394841 Journal of Biological Chemistry Creative Commons Attribution-Noncommercial-Share Alike http://creativecommons.org/licenses/by-nc-sa/4.0/ application/pdf American Society for Biochemistry and Molecular Biology (ASBMB) Walker via Courtney Crummett |
| spellingShingle | Wojtaszek, Jessica Lee, Chul-Jin D'Souza, Sanjay Victor Minesinger, Brenda Kim, Hyungjin D'Andrea, Alan D. Walker, Graham C. Zhou, Pei Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title | Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title_full | Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title_fullStr | Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title_full_unstemmed | Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title_short | Structural Basis of Rev1-mediated Assembly of a Quaternary Vertebrate Translesion Polymerase Complex Consisting of Rev1, Heterodimeric Polymerase (Pol) κ, and Pol ζ |
| title_sort | structural basis of rev1 mediated assembly of a quaternary vertebrate translesion polymerase complex consisting of rev1 heterodimeric polymerase pol κ and pol ζ |
| url | http://hdl.handle.net/1721.1/86395 https://orcid.org/0000-0001-7243-8261 |
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