Browse
Submit Data
Databases
API
Help

Dataset Information

27 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

UBC13-Mediated Ubiquitin Signaling Promotes Removal of Blocking Adducts from DNA Double-Strand Breaks.

ABSTRACT: Chemical modifications and adducts at DNA double-strand break (DSB) ends must be cleaned before re-joining by non-homologous end-joining (NHEJ). MRE11 nuclease is essential for efficient removal of Topoisomerase II (TOP2)-DNA adducts from TOP2 poison-induced DSBs. However, mechanisms in MRE11 recruitment to DSB sites in G₁ phase remain poorly understood. Here, we report that TOP2-DNA adducts are expeditiously removed through UBC13-mediated polyubiquitination, which promotes DSB resection in G₂ phase. We found that this ubiquitin signaling is required for efficient recruitment of MRE11 onto DSB sites in G₁ by facilitating localization of RAP80 and BRCA1 to DSB sites and complex formation between BRCA1 and MRE11 at DSB sites. UBC13 and MRE11 are dispensable for restriction-enzyme-induced "clean" DSBs repair but responsible for over 50% and 70% of NHEJ-dependent repair of γ-ray-induced "dirty" DSBs, respectively. In conclusion, ubiquitin signaling promotes nucleolytic removal of DSB blocking adducts by MRE11 before NHEJ.

SUBMITTER: Akagawa R

PROVIDER: S-EPMC7155233 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

The ubiquitin specific protease USP34 promotes ubiquitin signaling at DNA double-strand breaks.

Project description:Ubiquitylation plays key roles in DNA damage signal transduction. The current model envisions that lysine63-linked ubiquitin chains, via the concerted action of E3 ubiquitin ligases RNF8-RNF168, are built at DNA double-strand breaks (DSBs) to effectively assemble DNA damage-repair factors for proper checkpoint control and DNA repair. We found that RNF168 is a short-lived protein that is stabilized by the deubiquitylating enzyme USP34 in response to DNA damage. In the absence of USP34, RNF168 is rapidly degraded, resulting in attenuated DSB-associated ubiquitylation, defective recruitment of BRCA1 and 53BP1 and compromised cell survival after ionizing radiation. We propose that USP34 promotes a feed-forward loop to enforce ubiquitin signaling at DSBs and highlight critical roles of ubiquitin dynamics in genome stability maintenance.

| S-EPMC3794584 | biostudies-literature

The ubiquitin landscape at DNA double-strand breaks.

Project description:The intimate relationship between DNA double-strand break (DSB) repair and cancer susceptibility has sparked profound interest in how transactions on DNA and chromatin surrounding DNA damage influence genome integrity. Recent evidence implicates a substantial commitment of the cellular DNA damage response machinery to the synthesis, recognition, and hydrolysis of ubiquitin chains at DNA damage sites. In this review, we propose that, in order to accommodate parallel processes involved in DSB repair and checkpoint signaling, DSB-associated ubiquitin structures must be nonuniform, using different linkages for distinct functional outputs. We highlight recent advances in the study of nondegradative ubiquitin signaling at DSBs, and discuss how recognition of different ubiquitin structures may influence DNA damage responses.

| S-EPMC2779242 | biostudies-literature

WRN helicase promotes repair of DNA double-strand breaks caused by aberrant mismatch repair of chromium-DNA adducts.

Project description:Recent studies in yeast have found that processing of DNA double-strand breaks (DSB) for recombination repair involves Sgs1 helicase. Human cells have five Sgs1 homologues with unknown selectivity and significance for repair of different DSB types. Here we examined the importance of WRN helicase in repair of G(2)-specific DSB caused by abnormal mismatch repair (MMR) of ternary Cr-DNA adducts. We found that Cr(VI) induced a rapid dispersal of WRN from the nucleolus resulting in its prolonged retention in the nucleoplasm. The loss of MSH2 or MLH1 MMR proteins abolished the long-term but not the initial WRN relocalization. WRN-deficient fibroblasts were hypersensitive to Cr(VI)-induced clonogenic death and contained high levels of persistent DSB detected by gamma-H2AX/53BP1 foci and pulsed-field gel electrophoresis. WRN was involved in recombination repair of Cr-induced DNA damage, as evidenced by WRN-RAD51 colocalization and defective formation of RAD51 foci in the absence of WRN. The accumulation of unrepaired DSB in WRN-depleted cells was rescued by the inactivation of MMR, indicating that MMR-generated DSB were a key substrate for WRN action in Cr(VI)-treated cells. Competition for the limited amounts of WRN in primary cells between G(2) processes of telomere rebuilding and recombinational repair is expected to increase persistence of Cr-induced DSB and may cause telomeric abnormalities in tissues of chronically chromate-exposed workers. Our work provides the first demonstration of the major importance of WRN in repair of a specific class of DSB in human cells.

| S-EPMC3226781 | biostudies-literature

The Rap80-BRCC36 de-ubiquitinating enzyme complex antagonizes RNF8-Ubc13-dependent ubiquitination events at DNA double strand breaks.

Project description:DNA double strand breaks (DSBs) initiate reversible cellular checkpoint and repair activities. Whereas many of the activating events at DSBs have recently been elucidated, the mechanisms used to terminate responses at these sites are largely undefined. Here we report a pathway required to reverse RNF8-Ubc13 dependent ubiquitination events on chromatin flanking DSBs. Inhibition of the Rap80-BRCC36 de-ubiquitinating enzyme complex partially restored DSB-associated ubiquitin levels following RNF8 knockdown or proteasome inhibition. Similarly, BRCC36 knockdown or expression of a BRCC36 de-ubiquitinating enzyme-inactive mutant rescued both 53BP1 recruitment to DSBs and ionizing radiation-induced gammaH2AX ubiquitination following RNF8 depletion, and mitigated ionizing radiation sensitivity resulting from RNF8 deficiency. Thus, concomitant and opposing RNF8-Ubc13 ubiquitin ligase and Rap80-BRCC36 ubiquitin hydrolysis activities are responsible for determining steady-state ubiquitin levels at DNA DSBs. These findings reveal a Rap80-BRCC36 dependent pathway that is required for appropriate DSB recruitment and repair responses.

| S-EPMC2651241 | biostudies-literature

SCF ubiquitin E3 ligase regulates DNA double-strand breaks in early meiotic recombination.

Project description:Homeostasis of meiotic DNA double strand breaks (DSB) is critical for germline genome integrity and homologous recombination. Here we demonstrate an essential role for SKP1, a constitutive subunit of the SCF (SKP1-Cullin-F-box) ubiquitin E3 ligase, in early meiotic processes. SKP1 restrains accumulation of HORMAD1 and the pre-DSB complex (IHO1-REC114-MEI4) on the chromosome axis in meiotic germ cells. Loss of SKP1 prior to meiosis leads to aberrant localization of DSB repair proteins and a failure in synapsis initiation in meiosis of both males and females. Furthermore, SKP1 is crucial for sister chromatid cohesion during the pre-meiotic S-phase. Mechanistically, FBXO47, a meiosis-specific F-box protein, interacts with SKP1 and HORMAD1 and targets HORMAD1 for polyubiquitination and degradation in HEK293T cells. Our results support a model wherein the SCF ubiquitin E3 ligase prevents hyperactive DSB formation through proteasome-mediated degradation of HORMAD1 and subsequent modulation of the pre-DSB complex during meiosis.

| S-EPMC9122608 | biostudies-literature

Hedgehog signaling enables repair of ribosomal DNA double-strand breaks.

Project description:Ribosomal DNA (rDNA) consists of highly repeated sequences that are prone to incurring damage. Delays or failure of rDNA double-strand break (DSB) repair are deleterious, and can lead to rDNA transcriptional arrest, chromosomal translocations, genomic losses, and cell death. Here, we show that the zinc-finger transcription factor GLI1, a terminal effector of the Hedgehog (Hh) pathway, is required for the repair of rDNA DSBs. We found that GLI1 is activated in triple-negative breast cancer cells in response to ionizing radiation (IR) and localizes to rDNA sequences in response to both global DSBs generated by IR and site-specific DSBs in rDNA. Inhibiting GLI1 interferes with rDNA DSB repair and impacts RNA polymerase I activity and cell viability. Our findings tie Hh signaling to rDNA repair and this heretofore unknown function may be critically important in proliferating cancer cells.

| S-EPMC7544215 | biostudies-literature

RNF8 E3 Ubiquitin Ligase Stimulates Ubc13 E2 Conjugating Activity That Is Essential for DNA Double Strand Break Signaling and BRCA1 Tumor Suppressor Recruitment.

Project description:DNA double strand break (DSB) responses depend on the sequential actions of the E3 ubiquitin ligases RNF8 and RNF168 plus E2 ubiquitin-conjugating enzyme Ubc13 to specifically generate histone Lys-63-linked ubiquitin chains in DSB signaling. Here, we defined the activated RNF8-Ubc13?ubiquitin complex by x-ray crystallography and its functional solution conformations by x-ray scattering, as tested by separation-of-function mutations imaged in cells by immunofluorescence. The collective results show that the RING E3 RNF8 targets E2 Ubc13 to DSB sites and plays a critical role in damage signaling by stimulating polyubiquitination through modulating conformations of ubiquitin covalently linked to the Ubc13 active site. Structure-guided separation-of-function mutations show that the RNF8 E2 stimulating activity is essential for DSB signaling in mammalian cells and is necessary for downstream recruitment of 53BP1 and BRCA1. Chromatin-targeted RNF168 rescues 53BP1 recruitment involved in non-homologous end joining but not BRCA1 recruitment for homologous recombination. These findings suggest an allosteric approach to targeting the ubiquitin-docking cleft at the E2-E3 interface for possible interventions in cancer and chronic inflammation, and moreover, they establish an independent RNF8 role in BRCA1 recruitment.

| S-EPMC4850281 | biostudies-literature

Human RECQ5 helicase promotes repair of DNA double-strand breaks by synthesis-dependent strand annealing.

Project description:Most mitotic homologous recombination (HR) events proceed via a synthesis-dependent strand annealing mechanism to avoid crossing over, which may give rise to chromosomal rearrangements and loss of heterozygosity. The molecular mechanisms controlling HR sub-pathway choice are poorly understood. Here, we show that human RECQ5, a DNA helicase that can disrupt RAD51 nucleoprotein filaments, promotes formation of non-crossover products during DNA double-strand break-induced HR and counteracts the inhibitory effect of RAD51 on RAD52-mediated DNA annealing in vitro and in vivo. Moreover, we demonstrate that RECQ5 deficiency is associated with an increased occupancy of RAD51 at a double-strand break site, and it also causes an elevation of sister chromatid exchanges on inactivation of the Holliday junction dissolution pathway or on induction of a high load of DNA damage in the cell. Collectively, our findings suggest that RECQ5 acts during the post-synaptic phase of synthesis-dependent strand annealing to prevent formation of aberrant RAD51 filaments on the extended invading strand, thus limiting its channeling into potentially hazardous crossover pathway of HR.

| S-EPMC3936725 | biostudies-literature

GEMIN2 promotes accumulation of RAD51 at double-strand breaks in homologous recombination.

Project description:RAD51 is a key factor in homologous recombination (HR) and plays an essential role in cellular proliferation by repairing DNA damage during replication. The assembly of RAD51 at DNA damage is strictly controlled by RAD51 mediators, including BRCA1 and BRCA2. We found that human RAD51 directly binds GEMIN2/SIP1, a protein involved in spliceosome biogenesis. Biochemical analyses indicated that GEMIN2 enhances the RAD51-DNA complex formation by inhibiting RAD51 dissociation from DNA, and thereby stimulates RAD51-mediated homologous pairing. GEMIN2 also enhanced the RAD51-mediated strand exchange, when RPA was pre-bound to ssDNA before the addition of RAD51. To analyze the function of GEMIN2, we depleted GEMIN2 in the chicken DT40 line and in human cells. The loss of GEMIN2 reduced HR efficiency and resulted in a significant decrease in the number of RAD51 subnuclear foci, as observed in cells deficient in BRCA1 and BRCA2. These observations and our biochemical analyses reveal that GEMIN2 regulates HR as a novel RAD51 mediator.

| S-EPMC2926616 | biostudies-literature

RNF8 ubiquitinates RecQL4 and promotes its dissociation from DNA double strand breaks.

Project description:Ubiquitination-dependent DNA damage response (DDR) signals play a critical role in the cellular choice of DNA damage repair pathways. Human DNA helicase RecQL4 participates in DNA replication and repair, and loss of RecQL4 is associated with autosomal recessive genetic disorders characterized by genomic instability features. In an earlier study, RecQL4 was isolated as a stable complex that contained two ubiquitin ligases of the N-end rule (UBR1 and UBR2). However, it is unknown whether or not RecQL4 ubiquitination status is critical for its DNA repair function. Here, we report that RecQL4 directly interacts with RNF8 (a RING finger ubiquitin E3 ligase), and both co-localize at DNA double-strand break (DSB) sites. Our findings indicate that RNF8 ubiquitinates RecQL4 protein mainly at the lysine sites of 876, 1048, and 1101, thereby facilitating the dissociation of RecQL4 from DSB sites. RecQL4 mutant at ubiquitination sites had a significantly prolonged retention at DSBs, which hinders the recruitment of its direct downstream DSB repair proteins (CtIP & Ku80). Interestingly, reduced DSB repair capacity observed in RecQL4 depleted cells was restored only by the reconstitution of wild-type RecQL4, but not the ubiquitination mutant. Additionally, RecQL4 directly interacts with WRAP53β that is known to recruit RNF8 to DSBs and WRAP53β enhances the association of RecQL4 with RNF8. WRAP53β silencing resulted in a nearly diminished recruitment of RNF8 to DSBs and in a greatly attenuated dissociation of RecQL4 from the DSB sites. Collectively, our study demonstrates that the ubiquitination event mediated by RNF8 constitutes an essential component for RecQL4's function in DSB repair.

| S-EPMC7935965 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data