Non-enzymatic roles of human RAD51 at stalled replication forks

Mason, Jennifer M.; Chan, Yuen‐Ling; Weichselbaum, Ralph; Bishop, Douglas K.

doi:10.1101/359380

Cited by 17 publications

(24 citation statements)

References 63 publications

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“…Cells were harvested 72 hours post transfection. WT RAD51 or RAD51 II3A mutant was expressed from plasmid containing cDNA 13 .…”

Section: Methodsmentioning

confidence: 99%

“…4a ). Finally, we tested if RAD51 enzymatic activity is required for TERRA recruitment taking advantage of a mutation (RAD51 II3A) that retains DNA binding but not strand invasion activity 13 . In siRAD51 depleted cells, expression of wild type RAD51 from cDNA largely rescued TERRA colocalization with telomeres in contrast to RAD51 II3A ( Fig.…”

Section: Recombination Factors Promote Terra Association With Telomermentioning

confidence: 99%

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RAD51-dependent recruitment of TERRA lncRNA to telomeres through R-loops

Feretzaki

Pospíšilová

Fernandes

et al. 2020

Nature

147

138

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Telomeres which are present at the ends of eukaryotic chromosomes mediate genome stability and determine cellular lifespan 1 . Telomeric repeat containing RNAs (TERRA) are long noncoding RNAs transcribed from chromosome ends 2 , 3 , which regulate telomeric chromatin structure and telomere maintenance via telomerase and homology directed repair (HDR) 4 , 5 . The mechanisms by which TERRA is recruited to chromosome ends remain poorly defined. Here we develop a reporter system to dissect the underlying mechanisms and demonstrate that the UUAGGG-repeats of TERRA are both necessary and sufficient to target TERRA to chromosome ends. TERRA preferentially associates with short telomeres through the formation of telomeric DNA:RNA hybrid (R-loop) structures that can form in trans . Telomere association and R-loop formation triggers telomere fragility and is promoted by the RAD51 recombinase and its interacting partner BRCA2 but counteracted by RNA surveillance factors, RNaseH1 and TRF1. RAD51 physically interacts with TERRA and catalyzes R-loop formation with TERRA in vitro supporting a direct involvement of this DNA recombinase in TERRA recruitment by strand invasion. Together, our findings reveal a RAD51-dependent pathway that governs TERRA mediated R-loop formation post transcription providing a mechanism of how lncRNAs can be recruited to new loci in trans .

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“…Cells were harvested 72 hours post transfection. WT RAD51 or RAD51 II3A mutant was expressed from plasmid containing cDNA 13 .…”

Section: Methodsmentioning

confidence: 99%

Section: Recombination Factors Promote Terra Association With Telomermentioning

confidence: 99%

RAD51-dependent recruitment of TERRA lncRNA to telomeres through R-loops

Feretzaki

Pospíšilová

Fernandes

et al. 2020

Nature

147

138

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“…To distinguish the relative contributions of these RAD51 functions to MiDAS, we generated U2OS cells exogenously expressing RAD51 separation-of-function mutants, both of which have been shown to act in a dominant-negative manner. The RAD51 ATPase-dead K133R mutant forms highly stable RAD51 filaments which enhance RAD51-mediated protection of DNA but are deficient in HR (Mason et al, 2019;Morrison et al, 1999;Schlacher et al, 2011;Stark et al, 2002).…”

Section: The Role Of Rad51 In Fork Protection Promotes Midasmentioning

confidence: 99%

“…RAD51 also displays non-repair functions at stalled replication forks, in which RAD51 stimulates fork reversal and stabilises stalled forks by protecting against the nucleolytic degradation of nascent DNA (Mason et al, 2019;Morrison et al, 1999;Petermann et al, 2010;Schlacher et al, 2011;Schlacher et al, 2012;Wang et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

The RAD51 recombinase protects mitotic chromatin in human cells

Wassing

Saayman

Rampazzo

et al. 2020

Preprint

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The RAD51 recombinase plays critical roles in safeguarding genome integrity, which is fundamentally important for all living cells. While interphase functions of RAD51 in repairing broken DNA and protecting stalled replication forks are well characterised, its role in mitosis remains contentious. In this study, we show that RAD51 protects under-replicated DNA in mitotic human cells and, in this way, promotes mitotic DNA synthesis (MiDAS) and successful chromosome segregation. MiDAS was globally detectable irrespective of DNA damage and was promoted by de novo RAD51 recruitment, RAD51-mediated fork protection, and RAD51 phosphorylation by the key mitotic regulator Polo-like kinase 1. Importantly, acute inhibition of RAD51-promoted MiDAS led to mitotic DNA damage, delayed anaphase onset and induced centromere fragility, revealing a mechanism that prevents the satisfaction of the spindle assembly checkpoint when chromosomal replication remains incomplete. This study hence identifies an unexpected function of RAD51 in promoting the stability of mitotic chromatin.

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“…However, knockdown of RECQL1 in U2OS cells did not significantly reduce HR efficiency, as assessed using a green fluorescent protein (GFP)-based reporter assay ( Sharma et al, 2012 ). RECQL1, similar to RAD51, protects stalled replication forks from MRE11-dependent degradation ( Berti et al, 2013 ; Mason et al, 2019 ), suggesting that RECQL1 may play an indirect role in the protection and/or repair of one-ended DSBs at replication forks and is not directly required for strand invasion or homology search in HR at two-ended DSBs. Further studies are required to define the mechanism of RECQL1 in the repression of sister chromatid exchanges and its role in HR.…”

Section: Human Recq Helicasesmentioning

confidence: 99%

Human RecQ Helicases in DNA Double-Strand Break Repair

Davis

2021

Front. Cell Dev. Biol.

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RecQ DNA helicases are a conserved protein family found in bacteria, fungus, plants, and animals. These helicases play important roles in multiple cellular functions, including DNA replication, transcription, DNA repair, and telomere maintenance. Humans have five RecQ helicases: RECQL1, Bloom syndrome protein (BLM), Werner syndrome helicase (WRN), RECQL4, and RECQL5. Defects in BLM and WRN cause autosomal disorders: Bloom syndrome (BS) and Werner syndrome (WS), respectively. Mutations in RECQL4 are associated with three genetic disorders, Rothmund–Thomson syndrome (RTS), Baller–Gerold syndrome (BGS), and RAPADILINO syndrome. Although no genetic disorders have been reported due to loss of RECQL1 or RECQL5, dysfunction of either gene is associated with tumorigenesis. Multiple genetically independent pathways have evolved that mediate the repair of DNA double-strand break (DSB), and RecQ helicases play pivotal roles in each of them. The importance of DSB repair is supported by the observations that defective DSB repair can cause chromosomal aberrations, genomic instability, senescence, or cell death, which ultimately can lead to premature aging, neurodegeneration, or tumorigenesis. In this review, we will introduce the human RecQ helicase family, describe in detail their roles in DSB repair, and provide relevance between the dysfunction of RecQ helicases and human diseases.

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Non-enzymatic roles of human RAD51 at stalled replication forks

Cited by 17 publications

References 63 publications

RAD51-dependent recruitment of TERRA lncRNA to telomeres through R-loops

RAD51-dependent recruitment of TERRA lncRNA to telomeres through R-loops

The RAD51 recombinase protects mitotic chromatin in human cells

Human RecQ Helicases in DNA Double-Strand Break Repair

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