[Research Progress in the Roles of MRE11-RAD50-NBS1 Complex and Human Diseases].

DNA is susceptible to various factors in vitro and in vivo and experience different forms of damage,among which double-strand break(DSB)is a deleterious form.To maintain the stability of genetic information,organisms have developed multiple mechanisms to repair DNA damage.Among these mechanisms,homologous recombination(HR)is praised for the high accuracy.The MRE11-RAD50-NBS1(MRN)complex plays an important role in HR and is conserved across different species.The knowledge on the MRN complex mainly came from the previous studies in Saccharomyces cerevisiae and Caenorhabditis elegans,while studies in the last decades have revealed the role of mammalian MRN complex in DNA repair of higher animals.In this review,we first introduces the MRN complex regarding the composition,structure,and roles in HR.In addition,we discuss the human diseases such as ataxia-telangiectasia-like disorder,Nijmegen breakage syndrome,and Nijmegen breakage syndrome-like disorder that are caused by dysfunctions in the MRN complex.Furthermore,we summarize the mouse models established to study the clinical phenotypes of the above diseases.

Molecular mechanism of high glucose-induced mitochondrial DNA damage in retinal ganglion cells.

Mitochondrial DNA damage in retinal ganglion cells (RGCs) may be closely related to lesions of glaucoma. RGCs were cultured with different concentrations of glucose and grouped into 3 groups, namely normal control (NC) group, Low-Glu group, and High-Glu group. Cell viability was measured with cell counting kit-8, and cell apoptosis was measured using flow cytometry. The DNA damage was measured with comet assay, and the morphological changes of damaged mitochondria in RGCs were observed using TEM. Western blot analyzed the expression of MRE11, RAD50, and NBS1 protein. Cell viability of RGCs in Low-Glu and High-Glu groups were lower than that of NC group in 48 and 96 h. The cell apoptosis in NC group was 4.9%, the Low-Glu group was 12.2% and High-Glu group was 24.4%. The comet imaging showed that NC cells did not have tailings, but the low-Glu and high-Glu group cells had tailings, indicating that the DNA of RGCs had been damaged. TEM, mitochondrial membrane potential, ROS, mitochondrial oxygen consumption, and ATP content detection results showed that RGCs cultured with high glucose occurred mitochondrial morphology changes and dysfunction. MRE11, RAD50, and NBS1 protein expression associated with DNA damage repair pathway in High-Glu group declined compared with Low-Glu group. Mitochondrial DNA damage caused by high glucose will result in apoptosis of retinal ganglion cells in glaucoma.

Characterization of the signal transduction cascade for inflammatory gene expression in fibroblasts with ATM-ATR deficiencies after Ionizing radiation.

BACKGROUND AND PURPOSE: Ionizing radiation (IR) induces DNA double-strand breaks (DSBs), leading to micronuclei formation, which has emerged as a key mediator of inflammatory responses after IR. This study aimed to investigate the signaling cascade in inflammatory gene expression using fibroblasts harboring DNA damage response deficiency after exposure to IR. MATERIALS AND METHODS: Micronuclei formation was examined in human dermal fibroblasts derived from patients with deficiencies in ATM, ATR, MRE11, XLF, Artemis, or BRCA2 after IR. RNA-sequencing analysis was performed to assess gene expression, pathway mapping, and the balance of transcriptional activity using the transcription factor-based downstream gene expression mapping (TDEM) method developed in this study. RESULTS: Deficiencies in ATM, ATR, or MRE11 led to increased micronuclei formation after IR compared to normal cells. RNA-seq analysis revealed significant upregulation of inflammatory expression in cells deficient in ATM, ATR, or MRE11 following IR. Pathway mapping analysis identified the upregulation of RIG-I, MDA-5, IRF7, IL6, and interferon stimulated gene expression after IR. These changes were pronounced in cells deficient in ATM, ATR, or MRE11. TDEM analysis suggested the differential activation of STAT1/3-pathway between ATM and ATR deficiency. CONCLUSION: Enhanced micronuclei formation upon ATM, ATR, or MRE11 deficiency activated the cGAS/STING, RIG-I-MDA-5-IRF7-IL6 pathway, resulting in its downstream interferon stimulated gene expression following exposure to IR. Our study provides comprehensive information regarding the status of inflammation-related gene expression under DSB repair deficiency after IR. The generated dataset may be useful in developing functional biomarkers to accurately identify patients sensitive to radiotherapy.

Ccq1 restrains Mre11-mediated degradation to distinguish short telomeres from double-strand breaks.

Telomeres protect chromosome ends and are distinguished from DNA double-strand breaks (DSBs) by means of a specialized chromatin composed of DNA repeats bound by a multiprotein complex called shelterin. We investigated the role of telomere-associated proteins in establishing end-protection by studying viable mutants lacking these proteins. Mutants were studied using a Schizosaccharomyces pombe model system that induces cutting of a 'proto-telomere' bearing telomere repeats to rapidly form a new stable chromosomal end, in contrast to the rapid degradation of a control DSB. Cells lacking the telomere-associated proteins Taz1, Rap1, Poz1 or Rif1 formed a chromosome end that was stable. Surprisingly, cells lacking Ccq1, or impaired for recruiting Ccq1 to the telomere, converted the cleaved proto-telomere to a rapidly degraded DSB. Ccq1 recruits telomerase, establishes heterochromatin and affects DNA damage checkpoint activation; however, these functions were separable from protection of the new telomere by Ccq1. In cells lacking Ccq1, telomere degradation was greatly reduced by eliminating the nuclease activity of Mre11 (part of the Mre11-Rad50-Nbs1/Xrs2 DSB processing complex), and higher amounts of nuclease-deficient Mre11 associated with the new telomere. These results demonstrate a novel function for S. pombe Ccq1 to effect end-protection by restraining Mre11-dependent degradation of the DNA end.

Resection of DNA double-strand breaks activates Mre11-Rad50-Nbs1- and Rad9-Hus1-Rad1-dependent mechanisms that redundantly promote ATR checkpoint activation and end processing in Xenopus egg extracts.

Sensing and processing of DNA double-strand breaks (DSBs) are vital to genome stability. DSBs are primarily detected by the ATM checkpoint pathway, where the Mre11-Rad50-Nbs1 (MRN) complex serves as the DSB sensor. Subsequent DSB end resection activates the ATR checkpoint pathway, where replication protein A, MRN, and the Rad9-Hus1-Rad1 (9-1-1) clamp serve as the DNA structure sensors. ATR activation depends also on Topbp1, which is loaded onto DNA through multiple mechanisms. While different DNA structures elicit specific ATR-activation subpathways, the regulation and mechanisms of the ATR-activation subpathways are not fully understood. Using DNA substrates that mimic extensively resected DSBs, we show here that MRN and 9-1-1 redundantly stimulate Dna2-dependent long-range end resection and ATR activation in Xenopus egg extracts. MRN serves as the loading platform for ATM, which, in turn, stimulates Dna2- and Topbp1-loading. Nevertheless, MRN promotes Dna2-mediated end processing largely independently of ATM. 9-1-1 is dispensable for bulk Dna2 loading, and Topbp1 loading is interdependent with 9-1-1. ATR facilitates Mre11 phosphorylation and ATM dissociation. These data uncover that long-range end resection activates two redundant pathways that facilitate ATR checkpoint signaling and DNA processing in a vertebrate system.

Disordered regions mediate the interaction of p53 and MRE11.

The genome is frequently targeted by genotoxic agents, resulting in the formation of DNA scars. However, cells employ diverse repair mechanisms to restore DNA integrity. Among these processes, the Mre11-Rad50-Nbs1 complex detects double-strand breaks (DSBs) and recruits DNA damage response proteins such as ataxia-telangiectasia-mutated (ATM) kinase to DNA damage sites. ATM phosphorylates the transactivation domain (TAD) of the p53 tumor suppressor, which in turn regulates DNA repair, growth arrest, apoptosis, and senescence following DNA damage. The disordered glycine-arginine-rich (GAR) domain of double-strand break protein MRE11 (MRE11GAR) and its methylation are important for DSB repair, and localization to Promyelocytic leukemia nuclear bodies (PML-NBs). There is preliminary evidence that p53, PML protein, and MRE11 might co-localize and interact at DSB sites. To uncover the molecular details of these interactions, we aimed to identify the domains mediating the p53-MRE11 interaction and to elucidate the regulation of the p53-MRE11 interaction by post-translational modifications (PTMs) through a combination of biophysical techniques. We discovered that, in vitro, p53 binds directly to MRE11GAR mainly through p53TAD2 and that phosphorylation further enhances this interaction. Furthermore, we found that MRE11GAR methylation still allows for binding to p53. Overall, we demonstrated that p53 and MRE11 interaction is facilitated by disordered regions. We provide for the first time insight into the molecular details of the p53-MRE11 complex formation and elucidate potential regulatory mechanisms that will promote our understanding of the DNA damage response. Our findings suggest that PTMs regulate the p53-MRE11 interaction and subsequently their colocalization to PML-NBs upon DNA damage.

Bone Marrow Failure and Immunodeficiency Associated with Human RAD50 Variants.

PURPOSE: The MRE11-RAD50-NBN (MRN) complex plays a key role in recognizing and signaling DNA double-strand breaks. Pathogenic variants in NBN and MRE11 give rise to the autosomal-recessive diseases, Nijmegen breakage syndrome (NBS) and ataxia telangiectasia-like disorder, respectively. The clinical consequences of pathogenic variants in RAD50 are incompletely understood. We aimed to characterize a newly identified RAD50 deficiency/NBS-like disorder (NBSLD) patient with bone marrow failure and immunodeficiency. METHODS: We report on a girl with microcephaly, mental retardation, bird-like face, short stature, bone marrow failure and B-cell immunodeficiency. We searched for candidate gene by whole-exome sequencing and analyzed the cellular phenotype of patient-derived fibroblasts using immunoblotting, radiation sensitivity assays and lentiviral complementation experiments. RESULTS: Compound heterozygosity for two variants in the RAD50 gene (p.Arg83His and p.Glu485Ter) was identified in this patient. The expression of RAD50 protein and MRN complex formation was maintained in the cells derived from this patient. DNA damage-induced activation of the ATM kinase was markedly decreased, which was restored by the expression of wild-type (WT) RAD50. Radiosensitivity appeared inconspicuous in the patient-derived cell line as assessed by colony formation assay. The RAD50R83H missense substitution did not rescue the mitotic defect in complementation experiments using RAD50-deficient fibroblasts, whereas RAD50WT did. The RAD50E485X nonsense variant was associated with in-frame skipping of exon 10 (p.Glu485_545del). CONCLUSION: These findings indicate important roles of RAD50 in human bone marrow and immune cells. RAD50 deficiency/NBSLD can manifest as a distinct inborn error of immunity characterized by bone marrow failure and B-cell immunodeficiency.

Selective Killing of BRCA2-Deficient Ovarian Cancer Cells via MRE11 Blockade.

The MRE11 nuclease is essential during DNA damage recognition, homologous recombination, and replication. BRCA2 plays important roles during homologous recombination and replication. Here, we show that effecting an MRE11 blockade using a prototypical inhibitor (Mirin) induces synthetic lethality (SL) in BRCA2-deficient ovarian cancer cells, HeLa cells, and 3D spheroids compared to BRCA2-proficient controls. Increased cytotoxicity was associated with double-strand break accumulation, S-phase cell cycle arrest, and increased apoptosis. An in silico analysis revealed Mirin docking onto the active site of MRE11. While Mirin sensitises DT40 MRE11+/- cells to the Top1 poison SN-38, it does not sensitise nuclease-dead MRE11 cells to this compound confirming that Mirin specifically inhibits Mre11 nuclease activity. MRE11 knockdown reduced cell viability in BRCA2-deficient PEO1 cells but not in BRCA2-proficient PEO4 cells. In a Mirin-resistant model, we show the downregulation of 53BP1 and DNA repair upregulation, leading to resistance, including in in vivo xenograft models. In a clinical cohort of human ovarian tumours, low levels of BRCA2 expression with high levels of MRE11 co-expression were linked with worse progression-free survival (PFS) (p = 0.005) and overall survival (OS) (p = 0.001). We conclude that MRE11 is an attractive SL target, and the pharmaceutical development of MRE11 inhibitors for precision oncology therapeutics may be of clinical benefit.

Importance of Germline and Somatic Alterations in Human MRE11, RAD50, and NBN Genes Coding for MRN Complex.

The MRE11, RAD50, and NBN genes encode for the nuclear MRN protein complex, which senses the DNA double strand breaks and initiates the DNA repair. The MRN complex also participates in the activation of ATM kinase, which coordinates DNA repair with the p53-dependent cell cycle checkpoint arrest. Carriers of homozygous germline pathogenic variants in the MRN complex genes or compound heterozygotes develop phenotypically distinct rare autosomal recessive syndromes characterized by chromosomal instability and neurological symptoms. Heterozygous germline alterations in the MRN complex genes have been associated with a poorly-specified predisposition to various cancer types. Somatic alterations in the MRN complex genes may represent valuable predictive and prognostic biomarkers in cancer patients. MRN complex genes have been targeted in several next-generation sequencing panels for cancer and neurological disorders, but interpretation of the identified alterations is challenging due to the complexity of MRN complex function in the DNA damage response. In this review, we outline the structural characteristics of the MRE11, RAD50 and NBN proteins, the assembly and functions of the MRN complex from the perspective of clinical interpretation of germline and somatic alterations in the MRE11, RAD50 and NBN genes.

RBBP4 regulates the expression of the Mre11-Rad50-NBS1 (MRN) complex and promotes DNA double-strand break repair to mediate glioblastoma chemoradiotherapy resistance.

For treatment of glioblastoma (GBM), temozolomide (TMZ) and radiotherapy (RT) exert antitumor effects by inducing DNA double-strand breaks (DSBs), mainly via futile DNA mismatch repair (MMR) and inducing apoptosis. Here, we provide evidence that RBBP4 modulates glioblastoma resistance to chemotherapy and radiotherapy by recruiting transcription factors and epigenetic regulators that bind to their promoters to regulate the expression of the Mre11-Rad50-NBS1(MRN) complex and the level of DNA-DSB repair, which are closely associated with recovery from TMZ- and radiotherapy-induced DNA damage in U87MG and LN229 glioblastoma cells, which have negative MGMT expression. Disruption of RBBP4 induced GBM cell DNA damage and apoptosis in response to TMZ and radiotherapy and enhanced radiotherapy and chemotherapy sensitivity by the independent pathway of MGMT. These results displayed a possible chemo-radioresistant mechanism in MGMT negative GBM. In addition, the RBBP4-MRN complex regulation axis may provide an interesting target for developing therapy-sensitizing strategies for GBM.

The ends in sight: Mre11-Rad50-Nbs1 complex structures come into focus.

In this issue of Molecular Cell, Rotheneder et al.1 elucidate the eukaroytic Mre11-Rad50-Nbs1 (MRN) complex quaternary architecture, which together with cryo-EM structures of bacterial Mre11-Rad50-DNA complexes,2 resolves the basis for MRN assembly and its broad nuclease specificity regulating DNA double-strand break repair.

Deubiquitinase USP2 stabilizes the MRE11-RAD50-NBS1 complex at DNA double-strand break sites by counteracting the ubiquitination of NBS1.

The MRE11-RAD50-NBS1 (MRN) complex plays essential roles in the cellular response to DNA double-strand breaks (DSBs), which are the most cytotoxic DNA lesions, and is a target of various modifications and controls. Recently, lysine 48-linked ubiquitination of NBS1, resulting in premature disassembly of the MRN complex from DSB sites, was observed in cells lacking RECQL4 helicase activity. However, the role and control of this ubiquitination during the DSB response in cells with intact RECQL4 remain unknown. Here, we showed that USP2 counteracts this ubiquitination and stabilizes the MRN complex during the DSB response. By screening deubiquitinases that increase the stability of the MRN complex in RECQL4-deficient cells, USP2 was identified as a new deubiquitinase that acts at DSB sites to counteract NBS1 ubiquitination. We determined that USP2 is recruited to DSB sites in a manner dependent on ATM, a major checkpoint kinase against DSBs, and stably interacts with NBS1 and RECQL4 in immunoprecipitation experiments. Phosphorylation of two critical residues in the N terminus of USP2 by ATM is required for its recruitment to DSBs and its interaction with RECQL4. While inactivation of USP2 alone does not substantially influence the DSB response, we found that inactivation of USP2 and USP28, another deubiquitinase influencing NBS1 ubiquitination, results in premature disassembly of the MRN complex from DSB sites as well as defects in ATM activation and homologous recombination repair abilities. These results suggest that deubiquitinases counteracting NBS1 ubiquitination are essential for the stable maintenance of the MRN complex and proper cellular response to DSBs.

Cryo-EM structure of the Mre11-Rad50-Nbs1 complex reveals the molecular mechanism of scaffolding functions.

The DNA double-strand break repair complex Mre11-Rad50-Nbs1 (MRN) detects and nucleolytically processes DNA ends, activates the ATM kinase, and tethers DNA at break sites. How MRN can act both as nuclease and scaffold protein is not well understood. The cryo-EM structure of MRN from Chaetomium thermophilum reveals a 2:2:1 complex with a single Nbs1 wrapping around the autoinhibited Mre11 nuclease dimer. MRN has two DNA-binding modes, one ATP-dependent mode for loading onto DNA ends and one ATP-independent mode through Mre11's C terminus, suggesting how it may interact with DSBs and intact DNA. MRNs two 60-nm-long coiled-coil domains form a linear rod structure, the apex of which is assembled by the two joined zinc-hook motifs. Apices from two MRN complexes can further dimerize, forming 120-nm spanning MRN-MRN structures. Our results illustrate the architecture of MRN and suggest how it mechanistically integrates catalytic and tethering functions.

TRIM24 is critical for the cellular response to DNA double-strand breaks through regulating the recruitment of MRN complex.

The MRE11-RAD50-NBS1 (MRN) complex plays a crucial role in DNA double-strand breaks (DSBs) sensing and initiation of signaling cascades. However, the precise mechanisms by which the recruitment of MRN complex is regulated has yet to be elucidated. Here, we identified TRIpartite motif-containing protein 24 (TRIM24), a protein considered as an oncogene overexpressed in cancers, as a novel signaling molecule in response to DSBs. TRIM24 is essential for DSBs-induced recruitment of MRN complex and activation of downstream signaling. In the absence of TRIM24, MRN mediated DSBs repair is remarkably diminished. Mechanistically, TRIM24 is phosphorylated by ataxia-telangiectasia mutated (ATM) and then recruited to DSBs sites, facilitating the accumulation of the MRN components to chromatin. Depletion of TRIM24 sensitizes human hepatocellular carcinoma cells to cancer therapy agent-induced apoptosis and retards the tumor growth in a subcutaneous xenograft tumor mouse model. Together, our data reveal a novel function of TRIM24 in response to DSBs through regulating the MRN complex, which suggests that TRIM24 may be a potential therapeutic molecular target for tumor treatment.

An in-silico analysis to identify structural, functional and regulatory role of SNPs in hMRE11.

Meiotic recombination 11 (MRE11) is a component of the tri-molecular MRE11-RAD50-NBS1 (MRN) complex, which functions as an exonuclease and endonuclease which is involved in identifying, signalling, protecting and repairing double-strand breaks in DNA (DSBs). Ataxia-telangiectasia-like disorder (ATLD) 1 and Nijmegen breakage syndrome (NBS)-like disorder are MRE11 associated diseases. In the present study, we used an integrated computational approach to identify the most deleterious SNPs and their structural and functional impact on human MRE11. Five of the 68 observed non-synonymous SNP (nsSNPs; I162T, S273C, W210C, D311Y and R364L) should be worked on due to their strong possible pathogenicity and the risk of changing protein properties. All the nsSNPs were highly conserved and decrease the protein stability located in the MRE11 nuclease and MRE11 DNA binding presumed domain. R364L and I162T were predicted to be involved in post-translational modification (PTM) sites. Furthermore, we also analysed the regulatory effect of noncoding SNPs on MRE11 gene regulation in which 6 SNPs were found to affect gene regulation. All six noncoding SNPs predicted chromatin interactive site whereas only one SNP was noted its association with miRNA binding site which disrupts 5 miRNA conserved site. These findings help future studies to get more insights into the role of these variants in the alteration of the MRE11 function.Communicated by Ramaswamy H. Sarma.

TRIP13 Participates in Immediate-Early Sensing of DNA Strand Breaks and ATM Signaling Amplification through MRE11.

Thyroid hormone receptor-interacting protein 13 (TRIP13) participates in various regulatory steps related to the cell cycle, such as the mitotic spindle assembly checkpoint and meiotic recombination, possibly by interacting with members of the HORMA domain protein family. Recently, it was reported that TRIP13 could regulate the choice of the DNA repair pathway, i.e., homologous recombination (HR) or nonhomologous end-joining (NHEJ). However, TRIP13 is recruited to DNA damage sites within a few seconds after damage and may therefore have another function in DNA repair other than regulation of the pathway choice. Furthermore, the depletion of TRIP13 inhibited both HR and NHEJ, suggesting that TRIP13 plays other roles besides regulation of choice between HR and NHEJ. To explore the unidentified functions of TRIP13 in the DNA damage response, we investigated its genome-wide interaction partners in the context of DNA damage using quantitative proteomics with proximity labeling. We identified MRE11 as a novel interacting partner of TRIP13. TRIP13 controlled the recruitment of MDC1 to DNA damage sites by regulating the interaction between MDC1 and the MRN complex. Consistently, TRIP13 was involved in ATM signaling amplification. Our study provides new insight into the function of TRIP13 in immediate-early DNA damage sensing and ATM signaling activation.

POLθ prevents MRE11-NBS1-CtIP-dependent fork breakage in the absence of BRCA2/RAD51 by filling lagging-strand gaps.

POLθ promotes repair of DNA double-strand breaks (DSBs) resulting from collapsed forks in homologous recombination (HR) defective tumors. Inactivation of POLθ results in synthetic lethality with the loss of HR genes BRCA1/2, which induces under-replicated DNA accumulation. However, it is unclear whether POLθ-dependent DNA replication prevents HR-deficiency-associated lethality. Here, we isolated Xenopus laevis POLθ and showed that it processes stalled Okazaki fragments, directly visualized by electron microscopy, thereby suppressing ssDNA gaps accumulating on lagging strands in the absence of RAD51 and preventing fork reversal. Inhibition of POLθ DNA polymerase activity leaves fork gaps unprotected, enabling their cleavage by the MRE11-NBS1-CtIP endonuclease, which produces broken forks with asymmetric single-ended DSBs, hampering BRCA2-defective cell survival. These results reveal a POLθ-dependent genome protection function preventing stalled forks rupture and highlight possible resistance mechanisms to POLθ inhibitors.

Integrative Expression, Survival Analysis and Cellular miR-2909 Molecular Interplay in MRN Complex Check Point Sensor Genes (MRN-CSG) Involved in Breast Cancer.

BACKGROUND: Breast cancer, an emerging global challenge, is evidenced by recent studies of miRNAs involvement in DNA repair gene variants (MRE11, RAD50, and NBN as checkpoint sensor genes (CSG) - MRN-CSG). The identification of various mutations in MRN-CSG and their interactions with miRNAs is still not understood. The emerging studies of miR-2909 involvement in other cancers led us to explore its role as molecular mechanistic marker in breast cancer. MATERIALS AND METHODS: The genomic and proteomic data of MRN-CSG of breast cancer patients (8426 samples) was evaluated to identify the mutation types linked with the patient's survival rate. Additionally, molecular, 3D-structural and functional analysis was performed to identify miR-2909 as regulator of MRN-CSG. RESULTS: The genomic and proteomic data analysis shows genetic alterations with majority of missense mutations [RAD50 (0.7%), MRE11 (1.5%), and NBN (11%)], though with highest MRE11 mRNA expression in invasive ductal breast carcinoma as compared to other breast cancer types. The Kaplan-Meier survival curves suggest higher survival rate for unaltered groups as compared to the altered group. Network analysis and disease association of miR-2909 and MRN-CSG shows strong interactions with other partners. The molecular hybridization between miR-2909-RAD50 and miR-2909-MRE11 complexes showed thermodynamically stable structures. Further, argonaute protein, involved in RNA silencing, docking studies with miR-MRE11-mRNA and miR-RAD50-mRNA hybridized complexes showed strong binding affinity. CONCLUSION: The results suggest that miR-2909 forms strong thermodynamically stable molecular hybridized complexes with MRE11 and RAD50 mRNAs which further strongly interacts with argonaute protein to show potential molecular mechanistic role in breast cancer.

Crosstalk between SUMOylation and ubiquitylation controls DNA end resection by maintaining MRE11 homeostasis on chromatin.

DNA end resection is delicately regulated through various types of post-translational modifications to initiate homologous recombination, but the involvement of SUMOylation in this process remains incompletely understood. Here, we show that MRE11 requires SUMOylation to shield it from ubiquitin-mediated degradation when resecting damaged chromatin. Upon DSB induction, PIAS1 promotes MRE11 SUMOylation on chromatin to initiate DNA end resection. Then, MRE11 is deSUMOylated by SENP3 mainly after it has moved away from DSB sites. SENP3 deficiency results in MRE11 degradation failure and accumulation on chromatin, causing genome instability. We further show that cancer-related MRE11 mutants with impaired SUMOylation exhibit compromised DNA repair ability. Thus, we demonstrate that MRE11 SUMOylation in coordination with ubiquitylation is dynamically controlled by PIAS1 and SENP3 to facilitate DNA end resection and maintain genome stability.

METTL16 antagonizes MRE11-mediated DNA end resection and confers synthetic lethality to PARP inhibition in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers. Characterization of genetic alterations will improve our understanding and therapies for this disease. Here, we report that PDAC with elevated expression of METTL16, one of the 'writers' of RNA N6-methyladenosine modification, may benefit from poly-(ADP-ribose)-polymerase inhibitor (PARPi) treatment. Mechanistically, METTL16 interacts with MRE11 through RNA and this interaction inhibits MRE11's exonuclease activity in a methyltransferase-independent manner, thereby repressing DNA end resection. Upon DNA damage, ATM phosphorylates METTL16 resulting in a conformational change and autoinhibition of its RNA binding. This dissociates the METTL16-RNA-MRE11 complex and releases inhibition of MRE11. Concordantly, PDAC cells with high METTL16 expression show increased sensitivity to PARPi, especially when combined with gemcitabine. Thus, our findings reveal a role for METTL16 in homologous recombination repair and suggest that a combination of PARPi with gemcitabine could be an effective treatment strategy for PDAC with elevated METTL16 expression.