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Molecular basis of FIGNL1 in dissociating RAD51 from DNA and chromatin.

ABSTRACT: Maintaining genome integrity is an essential and challenging process. RAD51 recombinase, the central player of several crucial processes in repairing and protecting genome integrity, forms filaments on DNA. RAD51 filaments are tightly regulated. One of these regulators is FIGNL1, that prevents persistent RAD51 foci post-damage and genotoxic chromatin association in cells. The cryogenic electron microscopy structure of FIGNL1 in complex with RAD51 reveals that the FIGNL1 forms a non-planar hexamer and RAD51 N-terminus is enclosed in the FIGNL1 hexamer pore. Mutations in pore loop or catalytic residues of FIGNL1 render it defective in filament disassembly and are lethal in mouse embryonic stem cells. Our study reveals a unique mechanism for removing RAD51 from DNA and provides the molecular basis for FIGNL1 in maintaining genome stability.

SUBMITTER: Carver A

PROVIDER: S-EPMC11275795 | biostudies-literature | 2024 Jul

REPOSITORIES: biostudies-literature

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Molecular basis of FIGNL1 in dissociating RAD51 from DNA and chromatin.

Carver Alexander A Yu Tai-Yuan TY Yates Luke A LA White Travis T Wang Raymond R Lister Katie K Jasin Maria M Zhang Xiaodong X

bioRxiv : the preprint server for biology 20240716

Maintaining genome integrity is an essential and challenging process. RAD51 recombinase, the central player of several crucial processes in repairing and protecting genome integrity, forms filaments on DNA. RAD51 filaments are tightly regulated. One of these regulators is FIGNL1, that prevents persistent RAD51 foci post-damage and genotoxic chromatin association in cells. The cryogenic electron microscopy structure of FIGNL1 in complex with RAD51 reveals that the FIGNL1 forms a non-planar hexame ...[more]

PMID: 39071279

Similar Datasets

Molecular basis of FIGNL1 in dissociating RAD51 from DNA and chromatin.

Project description:Maintaining genome integrity is an essential and challenging process. RAD51 recombinase, the central component of several crucial processes in repairing DNA and protecting genome integrity, forms filaments on DNA, which are tightly regulated. One of these RAD51 regulators is FIGNL1 (fidgetin-like 1), which prevents RAD51 genotoxic chromatin association in normal cells and persistent RAD51 foci upon DNA damage. The cryogenic electron microscopy-imaged structure of FIGNL1 in complex with RAD51 reveals that FIGNL1 forms a nonplanar hexamer and encloses RAD51 N terminus in the FIGNL1 hexamer pore. Mutations in pore loop or catalytic residues of FIGNL1 render it defective in filament disassembly and are lethal in mouse embryonic stem cells. Our study reveals a distinct mechanism for removing RAD51 from bound substrates and provides the molecular basis for FIGNL1 in maintaining genome stability.

| S-EPMC7617353 | biostudies-literature

FIRRM cooperates with FIGNL1 to promote RAD51 disassembly during DNA repair.

Project description:Interstrand DNA cross-links (ICLs) represent complex lesions that compromise genomic stability. Several pathways have been involved in ICL repair, but the extent of factors involved in the resolution of ICL-induced DNA double-strand breaks (DSBs) remains poorly defined. Using CRISPR-based genomics, we identified FIGNL1 interacting regulator of recombination and mitosis (FIRRM) as a sensitizer of the ICL-inducing agent mafosfamide. Mechanistically, we showed that FIRRM, like its interactor Fidgetin like 1 (FIGNL1), contributes to the resolution of RAD51 foci at ICL-induced DSBs. While the stability of FIGNL1 and FIRRM is interdependent, expression of a mutant of FIRRM (∆WCF), which stabilizes the protein in the absence of FIGNL1, allows the resolution of RAD51 foci and cell survival, suggesting that FIRRM has FIGNL1-independent function during DNA repair. In line with this model, FIRRM binds preferentially single-stranded DNA in vitro, raising the possibility that it directly contributes to RAD51 disassembly by interacting with DNA. Together, our findings establish FIRRM as a promoting factor of ICL repair.

| S-EPMC10411901 | biostudies-literature

FLIP(C1orf112)-FIGNL1 complex regulates RAD51 chromatin association to promote viability after replication stress.

Project description:Homologous recombination (HR) plays critical roles in repairing lesions that arise during DNA replication and is thus essential for viability. RAD51 plays important roles during replication and HR, however, how RAD51 is regulated downstream of nucleofilament formation and how the varied RAD51 functions are regulated is not clear. We have investigated the protein c1orf112/FLIP that previously scored in genome-wide screens for mediators of DNA inter-strand crosslink (ICL) repair. Upon ICL agent exposure, FLIP loss leads to marked cell death, elevated chromosomal instability, increased micronuclei formation, altered cell cycle progression and increased DNA damage signaling. FLIP is recruited to damage foci and forms a complex with FIGNL1. Both proteins have epistatic roles in ICL repair, forming a stable complex. Mechanistically, FLIP loss leads to increased RAD51 amounts and foci on chromatin both with or without exogenous DNA damage, defective replication fork progression and reduced HR competency. We posit that FLIP is essential for limiting RAD51 levels on chromatin in the absence of damage and for RAD51 dissociation from nucleofilaments to properly complete HR. Failure to do so leads to replication slowing and inability to complete repair.

| S-EPMC10825145 | biostudies-literature

FIGNL1-FIRRM is essential for meiotic recombination and prevents DNA damage-independent RAD51 and DMC1 loading.

Project description:During meiosis, nucleoprotein filaments of the strand exchange proteins RAD51 and DMC1 are crucial for repairing SPO11-generated DNA double-strand breaks (DSBs) by homologous recombination (HR). A balanced activity of positive and negative RAD51/DMC1 regulators ensures proper recombination. Fidgetin-like 1 (FIGNL1) was previously shown to negatively regulate RAD51 in human cells. However, FIGNL1's role during meiotic recombination in mammals remains unknown. Here, we decipher the meiotic functions of FIGNL1 and FIGNL1 Interacting Regulator of Recombination and Mitosis (FIRRM) using male germline-specific conditional knock-out (cKO) mouse models. Both FIGNL1 and FIRRM are required for completing meiotic prophase in mouse spermatocytes. Despite efficient recruitment of DMC1 on ssDNA at meiotic DSB hotspots, the formation of late recombination intermediates is defective in Firrm cKO and Fignl1 cKO spermatocytes. Moreover, the FIGNL1-FIRRM complex limits RAD51 and DMC1 accumulation on intact chromatin, independently from the formation of SPO11-catalyzed DSBs. Purified human FIGNL1ΔN alters the RAD51/DMC1 nucleoprotein filament structure and inhibits strand invasion in vitro. Thus, this complex might regulate RAD51 and DMC1 association at sites of meiotic DSBs to promote proficient strand invasion and processing of recombination intermediates.

| S-EPMC11327267 | biostudies-literature

FIGNL1 AAA+ ATPase remodels RAD51 and DMC1 filaments in pre-meiotic DNA replication and meiotic recombination.

Project description:The formation of RAD51/DMC1 filaments on single-stranded (ss)DNAs essential for homology search and strand exchange in DNA double-strand break (DSB) repair is tightly regulated. FIGNL1 AAA+++ ATPase controls RAD51-mediated recombination in human cells. However, its role in gametogenesis remains unsolved. Here, we characterized a germ line-specific conditional knockout (cKO) mouse of FIGNL1. Fignl1 cKO male mice showed defective chromosome synapsis and impaired meiotic DSB repair with the accumulation of RAD51/DMC1 on meiotic chromosomes, supporting a positive role of FIGNL1 in homologous recombination at a post-assembly stage of RAD51/DMC1 filaments. Fignl1 cKO spermatocytes also accumulate RAD51/DMC1 on chromosomes in pre-meiotic S-phase. These RAD51/DMC1 assemblies are independent of meiotic DSB formation. We also showed that purified FIGNL1 dismantles RAD51 filament on double-stranded (ds)DNA as well as ssDNA. These results suggest an additional role of FIGNL1 in limiting the non-productive assembly of RAD51/DMC1 on native dsDNAs during pre-meiotic S-phase and meiotic prophase I.

| S-EPMC10611733 | biostudies-literature

Human RAD51 paralogue SWSAP1 fosters RAD51 filament by regulating the anti-recombinase FIGNL1 AAA+ ATPase.

Project description:RAD51 assembly on single-stranded (ss)DNAs is a crucial step in the homology-dependent repair of DNA damage for genomic stability. The formation of the RAD51 filament is promoted by various RAD51-interacting proteins including RAD51 paralogues. However, the mechanisms underlying the differential control of RAD51-filament dynamics by these factors remain largely unknown. Here, we report a role for the human RAD51 paralogue, SWSAP1, as a novel regulator of RAD51 assembly. Swsap1-deficient cells show defects in DNA damage-induced RAD51 assembly during both mitosis and meiosis. Defective RAD51 assembly in SWSAP1-depleted cells is suppressed by the depletion of FIGNL1, which binds to RAD51 as well as SWSAP1. Purified FIGNL1 promotes the dissociation of RAD51 from ssDNAs. The dismantling activity of FIGNL1 does not require its ATPase but depends on RAD51-binding. Purified SWSAP1 inhibits the RAD51-dismantling activity of FIGNL1. Taken together, our data suggest that SWSAP1 protects RAD51 filaments by antagonizing the anti-recombinase, FIGNL1.

| S-EPMC6440994 | biostudies-literature

FIGNL1 AAA+ ATPase remodels RAD51 and DMC1 filaments in meiotic DNA replication and recombination

Project description:The formation of RAD51/DMC1 filaments on single-stranded (ss)DNAs, which is essential for homology search and strand exchange in DNA double-strand break (DSB) repair and for the protection of stalled DNA replication forks, is tightly regulated in time and space. FIGNL1 AAA+++ ATPase plays positive and negative roles in the RAD51-mediated recombination in human cells. However, the role of FIGNL1 in gametogenesis remains unsolved. Here, we characterized a germ-line-specific conditional knockout (cKO) mouse of FIGNL1. The Fignl1 cKO male mice showed defective chromosome synapsis and impaired meiotic DSB repair with the accumulation of RAD51/DMC1 on chromosomes in mid-meiotic prophase I, supporting a role of FIGNL1 in a post-assembly stage of RAD51/DMC1 filaments. Fignl1 cKO spermatocytes accumulate RAD51 and DMC1 ensembles on chromosomes not only in early meiotic prophase I but also in meiotic S-phase. These RAD51/DMC1 assemblies are independent of meiotic DSB formation. Finally, we showed that purified FIGNL1 dismantles RAD51 filament on double-stranded (ds)DNA as well as ssDNA. These results suggest a critical role of FIGNL1 to limit the uncontrolled assembly of RAD51 and DMC1 on native dsDNAs during the meiotic S-phase and meiotic prophase I.

2023-03-31 | GSE227944 | GEO

The FIGNL1-interacting protein C1orf112 is synthetic lethal with PICH and mediates RAD51 retention on chromatin

Project description:Joint DNA molecules are natural by-products of DNA replication and repair. Persistent joint molecules give rise to ultrafine DNA bridges (UFBs) in mitosis, which compromise sister chromatid separation. The DNA translocase PICH (ERCC6L) plays a central role in UFB resolution. To better understand the genetic context rendering cells dependent on PICH, a genome-wide loss-of-function screen was performed to identify the genetic contexts in which cells become dependent on PICH. In addition to genes involved in DNA cohesion, centromere stability and DNA damage repair, we identified the uncharacterized protein C1orf112. We find that C1orf112 interacts with and stabilizes the AAA+ ATPase FIGNL1. Inactivation of either C1orf112 or FIGNL1 resulted in UFB formation, prolonged retention of RAD51 on chromatin, impaired replication fork dynamics, and consequently impaired genome maintenance. Combined, our data reveal that inactivation of C1orf112 or FIGNL1 dysregulates RAD51 dynamics at replication forks, resulting in DNA replication defects, and a dependency on PICH to preserve cell viability.

2024-08-08 | PXD035876 | Pride

Rad51 accumulation at sites of DNA damage and in postreplicative chromatin.

Project description:Rad51, a eukaryotic RecA homologue, plays a central role in homologous recombinational repair of DNA double-strand breaks (DSBs) in yeast and is conserved from yeast to human. Rad51 shows punctuate nuclear localization in human cells, called Rad51 foci, typically during the S phase (Tashiro, S., N. Kotomura, A. Shinohara, K. Tanaka, K. Ueda, and N. Kamada. 1996. Oncogene. 12:2165-2170). However, the topological relationships that exist in human S phase nuclei between Rad51 foci and damaged chromatin have not been studied thus far. Here, we report on ultraviolet microirradiation experiments of small nuclear areas and on whole cell ultraviolet C (UVC) irradiation experiments performed with a human fibroblast cell line. Before UV irradiation, nuclear DNA was sensitized by the incorporation of halogenated thymidine analogues. These experiments demonstrate the redistribution of Rad51 to the selectively damaged, labeled chromatin. Rad51 recruitment takes place from Rad51 foci scattered throughout the nucleus of nonirradiated cells in S phase. We also demonstrate the preferential association of Rad51 foci with postreplicative chromatin in contrast to replicating chromatin using a double labeling procedure with halogenated thymidine analogues. This finding supports a role of Rad51 in recombinational repair processes of DNA damage present in postreplicative chromatin.

| S-EPMC2180223 | biostudies-literature

RAD51 regulates eukaryotic chromatin motions in the absence of DNA damage.

Project description:In yeasts and higher eukaryotes, chromatin motions may be tuned to genomic functions, with transcriptional activation and the DNA damage response both leading to profound changes in chromatin dynamics. The RAD51 recombinase is a key mediator of chromatin mobility following DNA damage. As functions of RAD51 beyond DNA repair are being discovered, we asked whether RAD51 modulates chromatin dynamics in the absence of DNA damage and found that inhibition or depletion of RAD51 alters chromatin motions in undamaged cells. Inhibition of RAD51 increased nucleosome clustering. Predictions from polymer models are that chromatin clusters reduce chain mobility and, indeed, we measured reduced motion of individual chromatin loci in cells treated with a RAD51 inhibitor. This effect was conserved in mammalian cells, yeasts, and plant cells. In contrast, RAD51 depletion or inhibition increased global chromatin motions at the microscale. The results uncover a role for RAD51 in regulating local and global chromatin dynamics independently from DNA damage and highlight the importance of considering different physical scales when studying chromatin dynamics.

| S-EPMC11617103 | biostudies-literature

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