Biology subjects

Dovrat, D.

Publications and source records attributed to Dovrat, D..

3 recordsLinked to original sources

bioRxiv2026

Cruciform-forming AT/TA repeats are acted upon by structure selective endonucleases and Rad51 prior to repositioning to the nuclear periphery for repair

Structure-forming DNA repeats can pose a barrier to DNA replication and repair, creating chromosomal fragile sites. An AT/TA DNA repeat, derived from the Flex1 region of human common fragile site FRA16D, can form hairpin and cruciform structures which interfere with DNA replication. When inserted into the S. cerevisiae genome, the Flex1(AT)34 repeat stimulates chromosome deletions in a manner dependent on the Mus81-Mms4 nuclease and the SLX4 nuclease scaffold. It was previously found that hairpin forming CAG/CTG repeats move to the nuclear periphery to maintain genomic stability. Here, we show that a structure forming AT/TA repeat also relocalizes to the nuclear periphery in late S/G2 phase in a replication and length-dependent manner. In contrast to the CAG repeat, this shift in nuclear positioning is dependent on polySUMOylation and the activity of the Mus81-Mms4 nuclease. Processing by the Mre11 nuclease and Rad51-dependent strand exchange occurs prior to repositioning. Replication analysis indicates that the replisome likely bypasses the AT/TA repeat leaving behind a DNA structure that initiates relocation to the nuclear periphery. We conclude that AT/TA repeats form post-replicative DNA structures that are targeted for nuclease cleavage and require hairpin processing and repositioning to the nuclear periphery for homologous recombination-dependent repair.

genetics↗

bioRxiv2025

Replisome progression regulates R-loop mediated transcriptional repression

Maintaining cellular proliferation necessitates the synchronized activity of diverse molecular machineries operating in parallel on the genome. Wide-spread transcription and R-loop formation can interfere with genome duplication, causing transcription-replication conflicts (TRCs). Here, we use live-cell imaging for simultaneous monitoring of replication fork progression and transcription dynamics of an R-loop prone gene. While robust replisome progression through R-loops is observed in wild-type cells, it is impaired in RnaseH, Mph1 and Sen1 mutants. Intriguingly, we find that R-loop formation inhibits gene transcription, but this inhibition is reversed by replisome passage, demonstrating the dynamic crosstalk between replication, transcription and R-loops in a single cell cycle. Unexpectedly, we find that R-loops also have beneficial roles in reducing the density of RNAPII molecules on the gene, thereby preventing fork stalling upon high RNAPII occupancy. These findings illuminate that regulating R-loops and RNAPII density together is critical for preventing harmful TRCs.

cell biology↗

bioRxiv2020

Key histone chaperones have distinct roles in replisome progression and genomic stability

Replication-coupled (RC) nucleosome assembly is an essential process in eukaryotic cells in order to maintain chromatin structure during DNA replication. The deposition of newly synthesized H3/H4 histones during DNA replication is facilitated by specialized histone chaperones. Although the contribution of these histone chaperones to genomic stability has been thoroughly investigated, their effect on replisome progression is much less understood. By exploiting a time-lapse microscopy system for monitoring DNA replication in individual live cells, we examined how mutations in key histone chaperones including CAC1, RTT106, RTT109 and ASF1, affect replication fork progression. Our experiments revealed that mutations in CAC1 or RTT106 that directly deposit histones on the DNA, slowdown replication fork progression. In contrast, analysis of cells mutated in the intermediary ASF1 or RTT109 histone chaperones revealed that replisome progression is not affected. We found that mutations in histone chaperones including ASF1 and RTT109 lead to extended G2/M duration, elevated number of RPA foci and in some cases, increased spontaneous mutation rate. Our research suggests that histone chaperones have distinct roles in enabling high replisome progression and maintaining genome stability during cell cycle progression. Author SummaryHistone chaperones (HC) play key roles in maintaining the chromatin structure during DNA replication in eukaryotic cells. Despite extensive studies on HCs, little is known regarding their importance for replication fork progression during S-phase. Here, we utilized a live-cell imaging approach to measure the progression rates of single replication forks in individual yeast cells mutated in key histone chaperones. Using this approach, we show that mutations in CAC1 or RTT106 HCs that directly deposit histones on the DNA lead to slowdown of replication fork progression. In contrast, mutations in ASF1 or RTT109 HCs that transfers H3/H4 to CAC1 or RTT106, do not affect replisome progression but lead to post replication defects. Our results reveal distinct functions of HCs in replication fork progression and maintaining genome stability.

cell biology↗