RESUMO
Mesenchymal stem cells (MSC) are an appealing therapeutic cell type for many diseases. However, patients with poor health or advanced age often have MSCs with poor regenerative properties. A major limiter of MSC therapies is cellular senescence, which is marked by limited proliferation capability, diminished multipotency, and reduced regenerative properties. In this work, we explored the ability of applied mechanical forces to reduce cellular senescence in MSCs. Our studies revealed that mechanical conditioning caused a lasting enhancement in proliferation, overall cell culture expansion potential, multipotency, and a reduction of senescence in MSCs from aged donors. Mechanistic studies suggested that these functional enhancements were mediated by oxidative stress and DNA damage repair signaling with mechanical load altering the expression of proteins of the sirtuin pathway, the DNA damage repair protein ATM, and antioxidant proteins. In addition, our results suggest a biophysical mechanism in which mechanical stretch leads to improved recognition of damaged DNA in the nucleus. Analysis of the cells through RNA-seq and ATAC-seq, demonstrated that mechanical loading alters the cell's genetic landscape to cause broad shifts in transcriptomic patterns that related to senescence. Overall, our results demonstrate that mechanical conditioning can rejuvenate mesenchymal stem cells derived from aged patients and improve their potential as a therapeutic cell type.
RESUMO
Senataxin is an evolutionarily conserved RNA-DNA helicase involved in DNA repair and transcription termination that is associated with human neurodegenerative disorders. Here, we investigated whether Senataxin loss affects protein homeostasis based on previous work showing R-loop-driven accumulation of DNA damage and protein aggregates in human cells. We find that Senataxin loss results in the accumulation of insoluble proteins, including many factors known to be prone to aggregation in neurodegenerative disorders. These aggregates are located primarily in the nucleolus and are promoted by upregulation of non-coding RNAs expressed from the intergenic spacer region of ribosomal DNA. We also map sites of R-loop accumulation in human cells lacking Senataxin and find higher RNA-DNA hybrids within the ribosomal DNA, peri-centromeric regions, and other intergenic sites but not at annotated protein-coding genes. These findings indicate that Senataxin loss affects the solubility of the proteome through the regulation of transcription-dependent lesions in the nucleus and the nucleolus.
Assuntos
DNA Helicases , Enzimas Multifuncionais , RNA Helicases , RNA não Traduzido , Humanos , Nucléolo Celular/metabolismo , Nucléolo Celular/genética , Dano ao DNA , DNA Helicases/metabolismo , DNA Helicases/genética , DNA Ribossômico/genética , DNA Ribossômico/metabolismo , Enzimas Multifuncionais/metabolismo , Enzimas Multifuncionais/genética , Agregados Proteicos , Proteostase , Estruturas R-Loop/genética , RNA Helicases/metabolismo , RNA Helicases/genética , RNA não Traduzido/genética , RNA não Traduzido/metabolismoRESUMO
The ataxia telangiectasia mutated (ATM) protein kinase is a master regulator of the DNA damage response and also an important sensor of oxidative stress. Analysis of gene expression in ataxia-telangiectasia (A-T) patient brain tissue shows that large-scale transcriptional changes occur in patient cerebellum that correlate with the expression level and guanine-cytosine (GC) content of transcribed genes. In human neuron-like cells in culture, we map locations of poly(ADP-ribose) and RNA-DNA hybrid accumulation genome-wide with ATM inhibition and find that these marks also coincide with high transcription levels, active transcription histone marks, and high GC content. Antioxidant treatment reverses the accumulation of R-loops in transcribed regions, consistent with the central role of reactive oxygen species in promoting these lesions. Based on these results, we postulate that transcription-associated lesions accumulate in ATM-deficient cells and that the single-strand breaks and PARylation at these sites ultimately generate changes in transcription that compromise cerebellum function and lead to neurodegeneration over time in A-T patients.
Assuntos
Ataxia Telangiectasia , Poli Adenosina Difosfato Ribose , Humanos , RNA , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , DNA , Ataxia Telangiectasia/genética , Reparo do DNA , Dano ao DNA , Proteínas de Ciclo Celular/metabolismoRESUMO
Loss of the ATM protein kinase in humans results in Ataxia-telangiectasia, a disorder characterized by childhood-onset neurodegeneration of the cerebellum as well as cancer predisposition and immunodeficiency. Although many aspects of ATM function are well-understood, the mechanistic basis of the progressive cerebellar ataxia that occurs in patients is not. Here we review recent progress related to the role of ATM in neurons and the cerebellum that comes from many sources: animal models, post-mortem brain tissue samples, and human neurons in culture. These observations have revealed new insights into the consequences of ATM loss on DNA damage, gene expression, and immune signaling in the brain. Many results point to the importance of reactive oxygen species as well as single-strand DNA breaks in the progression of molecular events leading to neuronal dysfunction. In addition, innate immunity signaling pathways appear to play a critical role in ATM functions in microglia, responding to various forms of nucleic acid sensors and regulating survival of neurons and other cell types. Overall, the results lead to an updated view of transcriptional stress and DNA damage resulting from ATM loss that results in changes in gene expression as well as neuroinflammation that contribute to the cerebellar neurodegeneration observed in patients.
Assuntos
Ataxia Telangiectasia , Animais , Humanos , Criança , Ataxia Telangiectasia/genética , Cerebelo , Encéfalo , Dano ao DNA , DNA de Cadeia SimplesRESUMO
The ATM protein kinase is a master regulator of the DNA damage response and also an important sensor of oxidative stress. Analysis of gene expression in Ataxia-telangiectasia patient brain tissue shows that large-scale transcriptional changes occur in patient cerebellum that correlate with expression level and GC content of transcribed genes. In human neuron-like cells in culture we map locations of poly-ADP-ribose and RNA-DNA hybrid accumulation genome-wide with ATM inhibition and find that these marks also coincide with high transcription levels, active transcription histone marks, and high GC content. Antioxidant treatment reverses the accumulation of R-loops in transcribed regions, consistent with the central role of ROS in promoting these lesions. Based on these results we postulate that transcription-associated lesions accumulate in ATM-deficient cells and that the single-strand breaks and PARylation at these sites ultimately generate changes in transcription that compromise cerebellum function and lead to neurodegeneration over time in A-T patients.
RESUMO
The Mre11-Rad50-Nbs1 (MRN) complex recognizes and processes DNA double-strand breaks for homologous recombination by performing short-range removal of 5' strands. Endonucleolytic processing by MRN requires a stably bound protein at the break site-a role we postulate is played by DNA-dependent protein kinase (DNA-PK) in mammals. Here we interrogate sites of MRN-dependent processing by identifying sites of CtIP association and by sequencing DNA-PK-bound DNA fragments that are products of MRN cleavage. These intermediates are generated most efficiently when DNA-PK is catalytically blocked, yielding products within 200 bp of the break site, whereas DNA-PK products in the absence of kinase inhibition show greater dispersal. Use of light-activated Cas9 to induce breaks facilitates temporal resolution of DNA-PK and Mre11 binding, showing that both complexes bind to DNA ends before release of DNA-PK-bound products. These results support a sequential model of double-strand break repair involving collaborative interactions between homologous and non-homologous repair complexes.
Assuntos
Núcleo Celular , Quebras de DNA de Cadeia Dupla , Animais , Proteólise , Reparo do DNA , Proteína Quinase Ativada por DNA/genética , MamíferosRESUMO
Telomeres replicated by leading-strand synthesis lack the 3' overhang required for telomere protection. Surprisingly, resection of these blunt telomeres is initiated by the telomere-specific 5' exonuclease Apollo rather than the Mre11-Rad50-Nbs1 (MRN) complex, the nuclease that acts at DNA breaks. Without Apollo, leading-end telomeres undergo fusion, which, as demonstrated here, is mediated by alternative end joining. Here, we show that DNA-PK and TRF2 coordinate the repression of MRN at blunt mouse telomeres. DNA-PK represses an MRN-dependent long-range resection, while the endonuclease activity of MRN-CtIP, which could cleave DNA-PK off of blunt telomere ends, is inhibited in vitro and in vivo by the iDDR of TRF2. AlphaFold-Multimer predicts a conserved association of the iDDR with Rad50, potentially interfering with CtIP binding and MRN endonuclease activation. We propose that repression of MRN-mediated resection is a conserved aspect of telomere maintenance and represents an ancient feature of DNA-PK and the iDDR.
Assuntos
Quebras de DNA , Proteína Quinase Ativada por DNA , Animais , Camundongos , Endonucleases , Telômero , DNARESUMO
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.
Assuntos
Proteínas de Ligação a DNA , Proteínas Nucleares , Proteínas de Ligação a DNA/metabolismo , Proteína Homóloga a MRE11/genética , Proteínas Nucleares/metabolismo , Quebras de DNA de Cadeia Dupla , Dano ao DNA , DNARESUMO
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.
Assuntos
Carcinoma Ductal Pancreático , Proteína Homóloga a MRE11 , Metiltransferases , Neoplasias Pancreáticas , Adenosina Difosfato Ribose , Carcinoma Ductal Pancreático/tratamento farmacológico , DNA , Exonucleases/genética , Humanos , Proteína Homóloga a MRE11/genética , Metiltransferases/genética , Neoplasias Pancreáticas/tratamento farmacológico , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Poli(ADP-Ribose) Polimerases/genética , RNA , Mutações Sintéticas Letais , Neoplasias PancreáticasRESUMO
R loops occur frequently in genomes and contribute to fundamental biological processes at multiple levels. Consequently, understanding the molecular and cellular biology of R loops has become an emerging area of research. Here, it is shown that poly(ADP-ribose) polymerase-1 (PARP-1) can mediate the association of DDX18, a putative RNA helicase, with R loops thereby modulating R-loop homeostasis in endogenous R-loop-prone and DNA lesion regions. DDX18 depletion results in aberrant endogenous R-loop accumulation, which leads to DNA-replication defects. In addition, DDX18 depletion renders cells more sensitive to DNA-damaging agents and reduces RPA32 and RAD51 foci formation in response to irradiation. Notably, DDX18 depletion leads to γH2AX accumulation and genome instability, and RNase H1 overexpression rescues all the DNA-repair defects caused by DDX18 depletion. Taken together, these studies uncover a function of DDX18 in R-loop-mediated events and suggest a role for PARP-1 in mediating the binding of specific DDX-family proteins with R loops in cells.
Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases , Estruturas R-Loop , DNA , Dano ao DNA , Reparo do DNA , Instabilidade Genômica , HumanosRESUMO
Endonucleolytic cleavage of DNA ends by the human Mre11-Rad50-Nbs1 (MRN) complex occurs in a manner that is promoted by DNA-dependent protein kinase (DNA-PK). A method is described to isolate DNA-PK-bound fragments released from chromatin in human cells using a modified Gentle Lysis and Size Selection chromatin immunoprecipitation (GLASS-ChIP) protocol. This method, combined with real-time PCR or next-generation sequencing, can identify sites of MRN endonucleolytic cutting adjacent to DNA-PK binding sites in human cells.
Assuntos
Proteínas de Ligação a DNA , Proteínas Quinases , Imunoprecipitação da Cromatina , DNA/genética , DNA/metabolismo , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Proteínas Quinases/genéticaRESUMO
Endonucleolytic cleavage of DNA ends by the human Mre11-Rad50-Nbs1 (MRN) complex occurs in a manner that is promoted by DNA-dependent Protein Kinase (DNA-PK). A method is described to isolate DNA-PK-bound fragments released from chromatin in human cells using a modified Gentle Lysis and Size Selection chromatin immunoprecipitation (GLASS-ChIP) protocol. This method, combined with real-time PCR or next-generation sequencing, can identify sites of MRN endonucleolytic cutting adjacent to DNA-PK binding sites in human cells.
Assuntos
Proteínas de Ciclo Celular , Proteínas de Ligação a DNA , Hidrolases Anidrido Ácido/genética , Hidrolases Anidrido Ácido/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Imunoprecipitação da Cromatina , DNA/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Proteína Homóloga a MRE11/metabolismo , Proteínas Quinases/genéticaRESUMO
The protein kinase ataxia telangiectasia mutated (ATM) is a master regulator of double-strand DNA break (DSB) signalling and stress responses. For three decades, ATM has been investigated extensively to elucidate its roles in the DNA damage response (DDR) and in the pathogenesis of ataxia telangiectasia (A-T), a human neurodegenerative disease caused by loss of ATM. Although hundreds of proteins have been identified as ATM phosphorylation targets and many important roles for this kinase have been identified, it is still unclear how ATM deficiency leads to the early-onset cerebellar degeneration that is common in all individuals with A-T. Recent studies suggest the existence of links between ATM deficiency and other cerebellum-specific neurological disorders, as well as the existence of broader similarities with more common neurodegenerative disorders. In this Review, we discuss recent structural insights into ATM regulation, and possible aetiologies of A-T phenotypes, including reactive oxygen species, mitochondrial dysfunction, alterations in transcription, R-loop metabolism and alternative splicing, defects in cellular proteostasis and metabolism, and potential pathogenic roles for hyper-poly(ADP-ribosyl)ation.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Ataxia Telangiectasia/metabolismo , Doenças Neurodegenerativas/metabolismo , Ataxia Telangiectasia/genética , Ataxia Telangiectasia/patologia , Proteínas Mutadas de Ataxia Telangiectasia/química , Proteínas Mutadas de Ataxia Telangiectasia/deficiência , Reparo do DNA , Homeostase , Humanos , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/patologia , Oxirredução , Fosforilação , Poli(ADP-Ribose) Polimerases/metabolismo , RNA/metabolismoRESUMO
DNA double-strand breaks can be repaired through ligation-based pathways (non-homologous end-joining) or replication-based pathways (homologous recombination) in eukaryotic cells. The decisions that govern these outcomes are widely viewed as a competition between factors that recognize DNA ends and physically promote association of factors specific to each pathway, commonly known as 'pathway choice'. Here I review recent results in the literature and propose that this decision is better described as a sequential set of binding and end processing events, with non-homologous end joining as the first decision point. Physical association and co-localization of end resection factors with non-homologous end-joining factors suggests that ends are transferred between these complexes, thus the ultimate outcome is not the result of a competition but is more akin to a relay race that is determined by the efficiency of the initial end-joining event and the availability of activated DNA end-processing enzymes.
Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA por Junção de Extremidades , Animais , DNA/genética , Reparo do DNA por Junção de Extremidades/genética , Reparo do DNA/genética , Recombinação Homóloga , Mamíferos/genéticaRESUMO
The accumulation of unrepaired DNA lesions is associated with many pathological outcomes in humans, particularly in neurodegenerative diseases and in normal aging. Evidence supporting a causal role for DNA damage in the onset and progression of neurodegenerative disease has come from rare human patients with mutations in DNA damage response genes as well as from model organisms; however, the generality of this relationship in the normal population is unclear. In addition, the relevance of DNA damage in the context of proteotoxic stress-the widely accepted paradigm for pathology during neurodegeneration-is not well understood. Here, observations supporting intertwined roles of DNA damage and proteotoxicity in aging-related neurological outcomes are reviewed, with particular emphasis on recent insights into the relationships between DNA repair and autophagy, the ubiquitin proteasome system, formation of protein aggregates, poly-ADP-ribose polymerization, and transcription-driven DNA lesions.
Assuntos
Dano ao DNA , Reparo do DNA , Proteostase , Animais , Autofagia , DNA/metabolismo , Humanos , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/fisiopatologia , Poli Adenosina Difosfato Ribose/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina/metabolismoRESUMO
Loss of the ataxia-telangiectasia mutated (ATM) kinase causes cerebellum-specific neurodegeneration in humans. We previously demonstrated that deficiency in ATM activation via oxidative stress generates insoluble protein aggregates in human cells, reminiscent of protein dysfunction in common neurodegenerative disorders. Here, we show that this process is driven by poly-ADP-ribose polymerases (PARPs) and that the insoluble protein species arise from intrinsically disordered proteins associating with PAR-associated genomic sites in ATM-deficient cells. The lesions implicated in this process are single-strand DNA breaks dependent on reactive oxygen species, transcription, and R-loops. Human cells expressing Mre11 A-T-like disorder mutants also show PARP-dependent aggregation identical to ATM deficiency. Lastly, analysis of A-T patient cerebellum samples shows widespread protein aggregation as well as loss of proteins known to be critical in human spinocerebellar ataxias that is not observed in neocortex tissues. These results provide a hypothesis accounting for loss of protein integrity and cerebellum function in A-T.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/deficiência , Quebras de DNA de Cadeia Simples , Proteína Homóloga a MRE11/deficiência , Neocórtex/metabolismo , Poli ADP Ribosilação , Proteostase , Ataxias Espinocerebelares/metabolismo , Adulto , Linhagem Celular Tumoral , Feminino , Humanos , Masculino , Neocórtex/patologia , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/patologiaRESUMO
Homology-directed repair (HDR) safeguards DNA integrity under various forms of stress, but how HDR protects replicating genomes under extensive metabolic alterations remains unclear. Here, we report that besides stalling replication forks, inhibition of ribonucleotide reductase (RNR) triggers metabolic imbalance manifested by the accumulation of increased reactive oxygen species (ROS) in cell nuclei. This leads to a redox-sensitive activation of the ATM kinase followed by phosphorylation of the MRE11 nuclease, which in HDR-deficient settings degrades stalled replication forks. Intriguingly, nascent DNA degradation by the ROS-ATM-MRE11 cascade is also triggered by hypoxia, which elevates signaling-competent ROS and attenuates functional HDR without arresting replication forks. Under these conditions, MRE11 degrades daughter-strand DNA gaps, which accumulate behind active replisomes and attract error-prone DNA polymerases to escalate mutation rates. Thus, HDR safeguards replicating genomes against metabolic assaults by restraining mutagenic repair at aberrantly processed nascent DNA. These findings have implications for cancer evolution and tumor therapy.
Assuntos
Replicação do DNA , Genoma Humano , Metabolismo , Reparo de DNA por Recombinação , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteína BRCA2/deficiência , Proteína BRCA2/metabolismo , Hipóxia Celular , Linhagem Celular Tumoral , DNA/metabolismo , Humanos , Proteína Homóloga a MRE11/metabolismo , Modelos Biológicos , Mutação/genética , Neoplasias/genética , Neoplasias/patologia , Polimerização , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
DNA double-strand break (DSB) end resection initiates homologous recombination (HR) and is critical for genomic stability. DSB resection has been monitored indirectly in mammalian cells using detection of protein foci or BrdU foci formation, which is dependent on single-stranded DNA (ssDNA) products of resection. Here we describe a quantitative PCR (qPCR)-based assay to directly measure levels of ssDNA intermediates generated by resection at specific DSB sites in human cells, which is more quantitative and precise with respect to the extent and efficiency of resection compared with previous methods. This assay, excluding the time for making the stable cell line expressing the restriction enzyme AsiSI fused to the estrogen receptor hormone-binding domain (ER-AsiSI), can be completed within 3 days.
Assuntos
Quebras de DNA de Cadeia Dupla , Enzimas de Restrição do DNA/genética , Receptores de Estrogênio/genética , Reparo de DNA por Recombinação , Sítios de Ligação , Técnicas de Cultura de Células , Enzimas de Restrição do DNA/metabolismo , Instabilidade Genômica , Células HEK293 , Humanos , Receptores de Estrogênio/química , Receptores de Estrogênio/metabolismo , Proteínas Recombinantes de Fusão/metabolismoRESUMO
ATM kinase is a tumor suppressor and a master regulator of the DNA damage response. Most cancer-associated alterations to ATM are missense mutations at the PI3-kinase regulatory domain (PRD) or the kinase domain. Expression of kinase-dead (KD) ATM protein solely accelerates lymphomagenesis beyond ATM loss. To understand how PRD suppresses lymphomagenesis, we introduced the cancer-associated PRD mutation R3008H (R3016 in mouse) into mice. R3008H abrogated DNA damage- and oxidative stress-induced activation of ATM without consistently affecting ATM protein stability and recruitment. In contrast to the early embryonic lethality of AtmKD/KD mice, AtmR3016H (AtmR/R ) mice were viable, immunodeficient, and displayed spontaneous craniofacial abnormalities and delayed lymphomagenesis compared with Atm-/- controls. Mechanistically, R3008H rescued the tardy exchange of ATM-KD at DNA damage foci, indicating that PRD coordinates ATM activation with its exchange at DNA-breaks. Taken together, our results reveal a unique tumorigenesis profile for PRD mutations that is distinct from null or KD mutations. SIGNIFICANT: This study functionally characterizes the most common ATM missense mutation R3008H in cancer and identifies a unique role of PI3-kinase regulatory domain in ATM activation.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/genética , Dano ao DNA , Mutação , Neoplasias/genética , Animais , Ataxia Telangiectasia/genética , Ataxia Telangiectasia/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Células Cultivadas , Modelos Animais de Doenças , Embrião de Mamíferos/citologia , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Estimativa de Kaplan-Meier , Linfócitos/metabolismo , Linfócitos/patologia , Camundongos Knockout , Camundongos Transgênicos , Neoplasias/metabolismoRESUMO
The denitrosylase S-nitrosoglutathione reductase (GSNOR) has been suggested to sustain mitochondrial removal by autophagy (mitophagy), functionally linking S-nitrosylation to cell senescence and aging. In this study, we provide evidence that GSNOR is induced at the translational level in response to hydrogen peroxide and mitochondrial ROS. The use of selective pharmacological inhibitors and siRNA demonstrates that GSNOR induction is an event downstream of the redox-mediated activation of ATM, which in turn phosphorylates and activates CHK2 and p53 as intermediate players of this signaling cascade. The modulation of ATM/GSNOR axis, or the expression of a redox-insensitive ATM mutant influences cell sensitivity to nitrosative and oxidative stress, impairs mitophagy and affects cell survival. Remarkably, this interplay modulates T-cell activation, supporting the conclusion that GSNOR is a key molecular effector of the antioxidant function of ATM and providing new clues to comprehend the pleiotropic effects of ATM in the context of immune function.