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Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance

Many genome maintenance factors have multiple enzymatic activities. In most cases, how their distinct activities functionally relate with each other is unclear. Here we examined the conserved budding yeast Rad5 protein that has both ubiquitin ligase and DNA helicase activities. The Rad5 ubiquitin li...

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Autores principales: Choi, Koyi, Batke, Sabrina, Szakal, Barnabas, Lowther, Jonathan, Hao, Fanfan, Sarangi, Prabha, Branzei, Dana, Ulrich, Helle D., Zhao, Xiaolan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4357696/
https://www.ncbi.nlm.nih.gov/pubmed/25690888
http://dx.doi.org/10.1093/nar/gkv004
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author Choi, Koyi
Batke, Sabrina
Szakal, Barnabas
Lowther, Jonathan
Hao, Fanfan
Sarangi, Prabha
Branzei, Dana
Ulrich, Helle D.
Zhao, Xiaolan
author_facet Choi, Koyi
Batke, Sabrina
Szakal, Barnabas
Lowther, Jonathan
Hao, Fanfan
Sarangi, Prabha
Branzei, Dana
Ulrich, Helle D.
Zhao, Xiaolan
author_sort Choi, Koyi
collection PubMed
description Many genome maintenance factors have multiple enzymatic activities. In most cases, how their distinct activities functionally relate with each other is unclear. Here we examined the conserved budding yeast Rad5 protein that has both ubiquitin ligase and DNA helicase activities. The Rad5 ubiquitin ligase activity mediates PCNA poly-ubiquitination and subsequently recombination-based DNA lesion tolerance. Interestingly, the ligase domain is embedded in a larger helicase domain comprising seven consensus motifs. How features of the helicase domain influence ligase function is controversial. To clarify this issue, we use genetic, 2D gel and biochemical analyses and show that a Rad5 helicase motif important for ATP binding is also required for PCNA poly-ubiquitination and recombination-based lesion tolerance. We determine that this requirement is due to a previously unrecognized contribution of the motif to the PCNA and ubiquitination enzyme interaction, and not due to its canonical role in supporting helicase activity. We further show that Rad5′s helicase-mediated contribution to replication stress survival is separable from recombination. These findings delineate how two Rad5 enzymatic domains concertedly influence PCNA modification, and unveil their discrete contributions to stress tolerance.
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spelling pubmed-43576962015-03-20 Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance Choi, Koyi Batke, Sabrina Szakal, Barnabas Lowther, Jonathan Hao, Fanfan Sarangi, Prabha Branzei, Dana Ulrich, Helle D. Zhao, Xiaolan Nucleic Acids Res Genome Integrity, Repair and Replication Many genome maintenance factors have multiple enzymatic activities. In most cases, how their distinct activities functionally relate with each other is unclear. Here we examined the conserved budding yeast Rad5 protein that has both ubiquitin ligase and DNA helicase activities. The Rad5 ubiquitin ligase activity mediates PCNA poly-ubiquitination and subsequently recombination-based DNA lesion tolerance. Interestingly, the ligase domain is embedded in a larger helicase domain comprising seven consensus motifs. How features of the helicase domain influence ligase function is controversial. To clarify this issue, we use genetic, 2D gel and biochemical analyses and show that a Rad5 helicase motif important for ATP binding is also required for PCNA poly-ubiquitination and recombination-based lesion tolerance. We determine that this requirement is due to a previously unrecognized contribution of the motif to the PCNA and ubiquitination enzyme interaction, and not due to its canonical role in supporting helicase activity. We further show that Rad5′s helicase-mediated contribution to replication stress survival is separable from recombination. These findings delineate how two Rad5 enzymatic domains concertedly influence PCNA modification, and unveil their discrete contributions to stress tolerance. Oxford University Press 2015-03-11 2015-02-17 /pmc/articles/PMC4357696/ /pubmed/25690888 http://dx.doi.org/10.1093/nar/gkv004 Text en © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Genome Integrity, Repair and Replication
Choi, Koyi
Batke, Sabrina
Szakal, Barnabas
Lowther, Jonathan
Hao, Fanfan
Sarangi, Prabha
Branzei, Dana
Ulrich, Helle D.
Zhao, Xiaolan
Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title_full Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title_fullStr Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title_full_unstemmed Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title_short Concerted and differential actions of two enzymatic domains underlie Rad5 contributions to DNA damage tolerance
title_sort concerted and differential actions of two enzymatic domains underlie rad5 contributions to dna damage tolerance
topic Genome Integrity, Repair and Replication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4357696/
https://www.ncbi.nlm.nih.gov/pubmed/25690888
http://dx.doi.org/10.1093/nar/gkv004
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