Biology subjects

Kramarz, K.

Publications and source records attributed to Kramarz, K..

4 recordsLinked to original sources

bioRxiv2023

Replication stress response in fission yeast differentially depends on maintaining proper levels of Srs2 helicase and Rrp1, Rrp2 DNA translocases.

Homologous recombination is a key process that governs the stability of eukaryotic genomes during DNA replication and repair. Multiple auxiliary factors regulate the choice of homologous recombination pathway in response to different types of replication stress. Using Schizosaccharomyces pombe we have previously suggested the role of DNA translocases Rrp1 and Rrp2, together with Srs2 helicase, in the common synthesis dependent strand annealing sub-pathway of homologous recombination. Here we show that all three proteins are important for completion of replication after hydroxyurea exposure and provide data comparing the effect of overproduction of Srs2 with Rrp1 and Rrp2. Upregulation of Srs2 protein levels leads to enhanced replication stress, chromosome instability and viability loss, as previously reported for Rrp1 and Rrp2. Interestingly, our data suggests that dysregulation of Srs2, Rrp1 and Rrp2 protein levels differentially affects checkpoint response. Overproduction of Srs2 activates simultaneously DNA damage and replication stress response checkpoints, while cells overproducing Rrp1 mainly launch DNA damage checkpoint. Upregulation of Rrp2 primarily leads to replication stress response checkpoint activation. Overall, we propose that Srs2, Rrp1 and Rrp2 have important and independent functions for maintenance of distinct difficult to replicate regions of the genome.

cell biology↗

bioRxiv2023

SUMO protease and proteasome recruitment at the nuclear periphery differently affect replication dynamics at arrested forks.

Nuclear pores complexes (NPCs) are genome organizers, defining a particular nuclear compartment enriched for SUMO protease and proteasome activities, and acting as docking sites for DNA repair. In fission yeast, the anchorage of perturbed replication forks to NPCs is an integral part of the recombination-dependent replication restart mechanism (RDR) that resumes DNA synthesis at terminally dysfunctional forks. By mapping DNA polymerase usage, we report that SUMO protease Ulp1-associated NPCs ensure efficient initiation of restarted DNA synthesis, whereas proteasome-associated NPCs sustain the progression of restarted DNA polymerase. In contrast to Ulp1-dependent events, this last function occurs independently of SUMO chains formation. By analyzing the role of the nuclear basket, the nucleoplasmic extension of the NPC, we reveal that the activities of Ulp1 and the proteasome cannot compensate for each other and affect RDR dynamics in distinct ways. Our work probes the mechanisms by which the NPC environment ensures optimal RDR. Highlights[bullet] Ulp1-associated NPCs ensure efficient initiation of restarted DNA synthesis, in a SUMO chain-dependent manner [bullet]Proteasome-associated NPCs foster the progression of restarted DNA synthesis, in a SUMO chain-independent manner [bullet]The nucleoporin Nup60 promotes the spatial sequestration of Ulp1 at the nuclear periphery [bullet]Ulp1 and proteasome activities are differently required for optimal recombination-mediated fork restart.

molecular biology↗

bioRxiv2020

Rrp1 translocase and ubiquitin ligase activities restrict the genome destabilising effects of Rad51 in fission yeast

Rad51 is the key protein in homologous recombination DNA repair and has important roles during DNA replication. Auxiliary factors regulate Rad51 activity to facilitate productive, and prevent inappropriate, recombination that could lead to genome instability. Previous genetic analyses identified a function for Rrp1 (a member of the Rad5/16-like group of SWI2/SNF2 translocases) in counteracting Rad51 activity, shared with the Rad51 mediator Swi5-Sfr1 and the Srs2 anti-recombinase. Here, we show that Rrp1 overproduction alleviates the toxicity associated with excessive Rad51 activity in a manner dependent on Rrp1 ATPase domain. Purified Rrp1 binds to DNA and has a DNA-dependent ATPase activity. Importantly, Rrp1 directly interacts with Rad51 and removes it from double-stranded DNA, confirming that Rrp1 is a translocase capable of modulating Rad51 activity. Additionally, we demonstrate that Rrp1 possesses E3 ubiquitin ligase activity with Rad51 as a substrate, suggesting that Rrp1 regulates Rad51 in a multi-tiered fashion.

genetics↗

bioRxiv2019

S. pombe DNA translocases Rrp1 and Rrp2 have distinct roles at centromeres and telomeres that ensure genome stability.

Homologous recombination (HR) is a DNA repair mechanism that ensures, together with heterochromatin machinery, the proper replication, structure and function of telomeres and centromeres that is essential for the maintenance of genome integrity. Schizosaccharomyces pombe Rrp1 and Rrp2 participate in HR and are orthologues of Saccharomyces cerevisiae Uls1, a SWI2/SNF2 DNA translocase and SUMO-Targeted Ubiquitin Ligase. We show that Rrp1 or Rrp2 upregulation leads to chromosome instability and growth defects. These phenotypes depend on putative DNA translocase activities of Rrp1 and Rrp2. Either Rrp1 or Rrp2 overproduction results in a reduction in global histone levels, suggesting that Rrp1 and Rrp2 may modulate nucleosome dynamics. In addition we show that Rrp2, but not Rrp1, acts at telomeres. We propose that this role depends on the previously described interaction between Rrp2 and Top2. We conclude that Rrp1 and Rrp2 have important roles for centromere and telomere function and maintenance, contributing to the preservation of genome stability during vegetative cell growth.\n\nSUMMARY STATEMENTSchizosaccharomyces pombe DNA translocases Rrp1 and Rrp2 modulate centromere and telomere maintenance pathways and dysregulation of their activity leads to genome instability.

cell biology↗