Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics
The replication checkpoint is essential for accurate DNA replication and repair, and maintenance of genomic integrity when a cell is challenged with genotoxic stress. Several studies have defined the complement of proteins that change subcellular location in the budding yeast Saccharomyces cerevisia...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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eLife Sciences Publications Ltd
2023-06-01
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| Series: | eLife |
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| Online Access: | https://elifesciences.org/articles/82483 |
| _version_ | 1827932195493249024 |
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| author | Brandon Ho Ethan J Sanford Raphael Loll-Krippleber Nikko P Torres Marcus B Smolka Grant W Brown |
| author_facet | Brandon Ho Ethan J Sanford Raphael Loll-Krippleber Nikko P Torres Marcus B Smolka Grant W Brown |
| author_sort | Brandon Ho |
| collection | DOAJ |
| description | The replication checkpoint is essential for accurate DNA replication and repair, and maintenance of genomic integrity when a cell is challenged with genotoxic stress. Several studies have defined the complement of proteins that change subcellular location in the budding yeast Saccharomyces cerevisiae following chemically induced DNA replication stress using methyl methanesulfonate (MMS) or hydroxyurea (HU). How these protein movements are regulated remains largely unexplored. We find that the essential checkpoint kinases Mec1 and Rad53 are responsible for regulating the subcellular localization of 159 proteins during MMS-induced replication stress. Unexpectedly, Rad53 regulation of the localization of 52 proteins is independent of its known kinase activator Mec1, and in some scenarios independent of Tel1 or the mediator proteins Rad9 and Mrc1. We demonstrate that Rad53 is phosphorylated and active following MMS exposure in cells lacking Mec1 and Tel1. This noncanonical mode of Rad53 activation depends partly on the retrograde signaling transcription factor Rtg3, which also facilitates proper DNA replication dynamics. We conclude that there are biologically important modes of Rad53 protein kinase activation that respond to replication stress and operate in parallel to Mec1 and Tel1. |
| first_indexed | 2024-03-13T07:05:09Z |
| format | Article |
| id | doaj.art-237e6447f30d4f348e1780f7a1ef4b11 |
| institution | Directory Open Access Journal |
| issn | 2050-084X |
| language | English |
| last_indexed | 2024-03-13T07:05:09Z |
| publishDate | 2023-06-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj.art-237e6447f30d4f348e1780f7a1ef4b112023-06-06T11:55:27ZengeLife Sciences Publications LtdeLife2050-084X2023-06-011210.7554/eLife.82483Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamicsBrandon Ho0https://orcid.org/0000-0002-7566-3007Ethan J Sanford1Raphael Loll-Krippleber2Nikko P Torres3Marcus B Smolka4https://orcid.org/0000-0001-9952-2885Grant W Brown5https://orcid.org/0000-0002-9002-5003Department of Biochemistry, University of Toronto, Toronto, Canada; Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, CanadaThe Weill Institute for Cell & Molecular Biology, Cornell University, Ithaca, United StatesDepartment of Biochemistry, University of Toronto, Toronto, Canada; Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, CanadaDepartment of Biochemistry, University of Toronto, Toronto, Canada; Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, CanadaThe Weill Institute for Cell & Molecular Biology, Cornell University, Ithaca, United StatesDepartment of Biochemistry, University of Toronto, Toronto, Canada; Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, CanadaThe replication checkpoint is essential for accurate DNA replication and repair, and maintenance of genomic integrity when a cell is challenged with genotoxic stress. Several studies have defined the complement of proteins that change subcellular location in the budding yeast Saccharomyces cerevisiae following chemically induced DNA replication stress using methyl methanesulfonate (MMS) or hydroxyurea (HU). How these protein movements are regulated remains largely unexplored. We find that the essential checkpoint kinases Mec1 and Rad53 are responsible for regulating the subcellular localization of 159 proteins during MMS-induced replication stress. Unexpectedly, Rad53 regulation of the localization of 52 proteins is independent of its known kinase activator Mec1, and in some scenarios independent of Tel1 or the mediator proteins Rad9 and Mrc1. We demonstrate that Rad53 is phosphorylated and active following MMS exposure in cells lacking Mec1 and Tel1. This noncanonical mode of Rad53 activation depends partly on the retrograde signaling transcription factor Rtg3, which also facilitates proper DNA replication dynamics. We conclude that there are biologically important modes of Rad53 protein kinase activation that respond to replication stress and operate in parallel to Mec1 and Tel1.https://elifesciences.org/articles/82483DNA repairprotein localizationcell cycle checkpointprotein kinaseretrograde signaling |
| spellingShingle | Brandon Ho Ethan J Sanford Raphael Loll-Krippleber Nikko P Torres Marcus B Smolka Grant W Brown Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics eLife DNA repair protein localization cell cycle checkpoint protein kinase retrograde signaling |
| title | Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| title_full | Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| title_fullStr | Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| title_full_unstemmed | Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| title_short | Mec1-independent activation of the Rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| title_sort | mec1 independent activation of the rad53 checkpoint kinase revealed by quantitative analysis of protein localization dynamics |
| topic | DNA repair protein localization cell cycle checkpoint protein kinase retrograde signaling |
| url | https://elifesciences.org/articles/82483 |
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