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Deficiency of Polη in Saccharomyces cerevisiae reveals the impact of transcription on damage-induced cohesion.

ABSTRACT: The structural maintenance of chromosome (SMC) complex cohesin mediates sister chromatid cohesion established during replication, and damage-induced cohesion formed in response to DSBs post-replication. The translesion synthesis polymerase Polη is required for damage-induced cohesion through a hitherto unknown mechanism. Since Polη is functionally associated with transcription, and transcription triggers de novo cohesion in Schizosaccharomyces pombe, we hypothesized that transcription facilitates damage-induced cohesion in Saccharomyces cerevisiae. Here, we show dysregulated transcriptional profiles in the Polη null mutant (rad30Δ), where genes involved in chromatin assembly and positive transcription regulation were downregulated. In addition, chromatin association of RNA polymerase II was reduced at promoters and coding regions in rad30Δ compared to WT cells, while occupancy of the H2A.Z variant (Htz1) at promoters was increased in rad30Δ cells. Perturbing histone exchange at promoters inactivated damage-induced cohesion, similarly to deletion of the RAD30 gene. Conversely, altering regulation of transcription elongation suppressed the deficient damage-induced cohesion in rad30Δ cells. Furthermore, transcription inhibition negatively affected formation of damage-induced cohesion. These results indicate that the transcriptional deregulation of the Polη null mutant is connected with its reduced capacity to establish damage-induced cohesion. This also suggests a linkage between regulation of transcription and formation of damage-induced cohesion after replication.

SUBMITTER: Wu PS

PROVIDER: S-EPMC8454932 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Importance of Polη for damage-induced cohesion reveals differential regulation of cohesion establishment at the break site and genome-wide.

Project description:Genome integrity depends on correct chromosome segregation, which in turn relies on cohesion between sister chromatids from S phase until anaphase. S phase cohesion, together with DNA double-strand break (DSB) recruitment of cohesin and formation of damage-induced (DI) cohesion, has previously been shown to be required also for efficient postreplicative DSB repair. The budding yeast acetyltransferase Eco1 (Ctf7) is a common essential factor for S phase and DI-cohesion. The fission yeast Eco1 ortholog, Eso1, is expressed as a fusion protein with the translesion synthesis (TLS) polymerase Polη. The involvement of Eso1 in S phase cohesion was attributed to the Eco1 homologous part of the protein and bypass of UV-induced DNA lesions to the Polη part. Here we describe an additional novel function for budding yeast Polη, i.e. formation of postreplicative DI genome-wide cohesion. This is a unique Polη function not shared with other TLS polymerases. However, Polη deficient cells are DSB repair competent, as Polη is not required for cohesion locally at the DSB. This reveals differential regulation of DSB-proximal cohesion and DI genome-wide cohesion, and challenges the importance of the latter for DSB repair. Intriguingly, we found that specific inactivation of DI genome-wide cohesion increases chromosomal mis-segregation at the entrance of the next cell cycle, suggesting that S phase cohesion is not sufficient for correct chromosome segregation in the presence of DNA damage.

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Post-translational Regulation of DNA Polymerase ?, a Connection to Damage-Induced Cohesion in <i>Saccharomyces cerevisiae</i>.

Project description:Double-strand breaks that are induced postreplication trigger establishment of damage-induced cohesion in Saccharomyces cerevisiae, locally at the break site and genome-wide on undamaged chromosomes. The translesion synthesis polymerase, polymerase ?, is required for generation of damage-induced cohesion genome-wide. However, its precise role and regulation in this process is unclear. Here, we investigated the possibility that the cyclin-dependent kinase Cdc28 and the acetyltransferase Eco1 modulate polymerase ? activity. Through in vitro phosphorylation and structure modeling, we showed that polymerase ? is an attractive substrate for Cdc28 Mutation of the putative Cdc28-phosphorylation site Ser14 to Ala not only affected polymerase ? protein level, but also prevented generation of damage-induced cohesion in vivo We also demonstrated that Eco1 acetylated polymerase ? in vitro Certain nonacetylatable polymerase ? mutants showed reduced protein level, deficient nuclear accumulation, and increased ultraviolet irradiation sensitivity. In addition, we found that both Eco1 and subunits of the cohesin network are required for cell survival after ultraviolet irradiation. Our findings support functionally important Cdc28-mediated phosphorylation, as well as post-translational modifications of multiple lysine residues that modulate polymerase ? activity, and provide new insights into understanding the regulation of polymerase ? for damage-induced cohesion.

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Importance of Polη for damage-induced cohesion reveals differential regulation of cohesion establishment at the break site and genome-wide

Project description:Correct chromosome segregation requires that sister chromatids are held together by the protein complex cohesin, from S phase until anaphase. This S phase established cohesion is, together with DSB recruitment of cohesin and formation of damage induced (DI) cohesion, also important for repair of DSBs. Eco1 is a common essential factor for S phase and DI-cohesion. The fission yeast Eco1ortholog, Eso1, is important both for S phase cohesion and for bypass of UV induced lesions, and is expressed as a fusion protein with Polη. The cohesion function has been attributed solely to Eso1 and the lesion bypass function to the Polη part of the protein. As we found the interaction between the two proteins intriguing we decided to look for a functional connection also in budding yeast. Indeed, despite being dispensable for S phase cohesion, budding yeast Polη is required for formation of DI genome-wide cohesion. However, Polη deficient cells are DSB repair competent, revealing differential regulation of DI-cohesion at the break and genome-wide. This finding challenges the importance of DI genome-wide cohesion for DSB repair, and based on our findings we suggest that S phase cohesion is not sufficient for correct chromosome segregation in the presence of DNA damage. Whole Genome binding of G2 expressed Scc1 in the presence and absence of Rad30.

2013-01-15 | E-GEOD-42655 | biostudies-arrayexpress

Importance of Polη for damage-induced cohesion reveals differential regulation of cohesion establishment at the break site and genome-wide

2013-01-15 | GSE42655 | GEO

DNA damage induces Yap5-dependent transcription of ECO1/CTF7 in Saccharomyces cerevisiae.

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