Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The non-homologous end-joining protein Nej1p is a target of the DNA damage checkpoint.

ABSTRACT: DNA double-strand breaks (DSBs), which are generated by ionizing radiation (IR) and a range of other DNA damaging agents, are repaired by homologous recombination (HR) or non-homologous end-joining (NHEJ). Previous studies have shown that NHEJ in Saccharomyces cerevisiae requires the Yku70p-Yku80p heterodimer and a complex consisting of DNA Ligase IV, Lif1p and Nej1p. Here, we report that Nej1p is phosphorylated in response to DNA damage in a manner that relies on the DNA damage checkpoint kinases Mec1p, Rad53p and Dun1p. By using a mutational approach, we have identified a consensus Dun1p phosphorylation site in Nej1p, and mutation of conserved serine residues within it leads to decreased NHEJ efficiency. These data, together with previous findings that Rad55p--a protein involved in HR--is phosphorylated analogously, point to there being a broad signalling network connecting DNA damage checkpoint responses with the regulation of DNA DSB repair activities.

SUBMITTER: Ahnesorg P

PROVIDER: S-EPMC3338008 | biostudies-literature | 2007 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

The non-homologous end-joining protein Nej1p is a target of the DNA damage checkpoint.

Ahnesorg Peter P Jackson Stephen P SP

DNA repair 20061113 2

DNA double-strand breaks (DSBs), which are generated by ionizing radiation (IR) and a range of other DNA damaging agents, are repaired by homologous recombination (HR) or non-homologous end-joining (NHEJ). Previous studies have shown that NHEJ in Saccharomyces cerevisiae requires the Yku70p-Yku80p heterodimer and a complex consisting of DNA Ligase IV, Lif1p and Nej1p. Here, we report that Nej1p is phosphorylated in response to DNA damage in a manner that relies on the DNA damage checkpoint kinas ...[more]

PMID: 17101301

Similar Datasets

The checkpoint clamp protein Rad9 facilitates DNA-end resection and prevents alternative non-homologous end joining.

Project description:DNA damage activates the cell cycle checkpoint to regulate cell cycle progression. The checkpoint clamp (Rad9-Hus1-Rad1 complex) is recruited to damage sites, and is required for checkpoint activation. While functions of the checkpoint clamp in checkpoint activation have been well studied, its functions in DNA repair regulation remain elusive. Here we show that Rad9 is required for efficient homologous recombination (HR), and facilitates DNA-end resection. The role of Rad9 in homologous recombination is independent of its function in checkpoint activation, and this function is important for preventing alternative non-homologous end joining (altNHEJ). These findings reveal novel function of the checkpoint clamp in HR.

| S-EPMC4614839 | biostudies-other

MRI Is a DNA Damage Response Adaptor during Classical Non-homologous End Joining.

Project description:The modulator of retrovirus infection (MRI or CYREN) is a 30-kDa protein with a conserved N-terminal Ku-binding motif (KBM) and a C-terminal XLF-like motif (XLM). We show that MRI is intrinsically disordered and interacts with many DNA damage response (DDR) proteins, including the kinases ataxia telangiectasia mutated (ATM) and DNA-PKcs and the classical non-homologous end joining (cNHEJ) factors Ku70, Ku80, XRCC4, XLF, PAXX, and XRCC4. MRI forms large multimeric complexes that depend on its N and C termini and localizes to DNA double-strand breaks (DSBs), where it promotes the retention of DDR factors. Mice deficient in MRI and XLF exhibit embryonic lethality at a stage similar to those deficient in the core cNHEJ factors XRCC4 or DNA ligase IV. Moreover, MRI is required for cNHEJ-mediated DSB repair in XLF-deficient lymphocytes. We propose that MRI is an adaptor that, through multivalent interactions, increases the avidity of DDR factors to DSB-associated chromatin to promote cNHEJ.

| S-EPMC6083883 | biostudies-literature

Ku70 and non-homologous end joining protect testicular cells from DNA damage.

Project description:Spermatogenesis is a complex process that generates haploid germ cells or spores and implements meiosis, a succession of two special cell divisions that are required for homologous chromosome segregation. During prophase to the first meiotic division, homologous recombination (HR) repairs Spo11-dependent DNA double-strand breaks (DSBs) in the presence of telomere movements to allow for chromosome pairing and segregation at the meiosis I division. In contrast to HR, non-homologous end joining (NHEJ), the major DSB repair mechanism during the G1 cell cycle phase, is downregulated during early meiotic prophase. At somatic mammalian telomeres, the NHEJ factor Ku70/80 inhibits HR, as does the Rap1 component of the shelterin complex. Here, we investigated the role of Ku70 and Rap1 in meiotic telomere redistribution and genome protection in spermatogenesis by studying single and double knockout mice. Ku70(-/-) mice display reduced testis size and compromised spermatogenesis, whereas meiotic telomere dynamics and chromosomal bouquet formation occurred normally in Ku70(-/-) and Ku70(-/-)Rap1(?/?) knockout spermatocytes. Elevated mid-preleptotene frequencies were associated with significantly increased DNA damage in Ku-deficient B spermatogonia, and in differentiated Sertoli cells. Significantly elevated levels of ?H2AX foci in Ku70(-/-) diplotene spermatocytes suggest compromised progression of DNA repair at a subset of DSBs. This might explain the elevated meiotic metaphase apoptosis that is present in Ku70-deficient stage XII testis tubules, indicating spindle assembly checkpoint activation. In summary, our data indicate that Ku70 is important for repairing DSBs in somatic cells and in late spermatocytes of the testis, thereby assuring the fidelity of spermatogenesis.

| S-EPMC3711201 | biostudies-other

PHF6 promotes non-homologous end joining and G2 checkpoint recovery.

Project description:The cellular response to DNA breaks is influenced by chromatin compaction. To identify chromatin regulators involved in the DNA damage response, we screened for genes that affect recovery following DNA damage using an RNAi library of chromatin regulators. We identified genes involved in chromatin remodeling, sister chromatid cohesion, and histone acetylation not previously associated with checkpoint recovery. Among these is the PHD finger protein 6 (PHF6), a gene mutated in Börjeson-Forssman-Lehmann syndrome and leukemic cancers. We find that loss of PHF6 dramatically compromises checkpoint recovery in G2 phase cells. Moreover, PHF6 is rapidly recruited to sites of DNA lesions in a PARP-dependent manner and required for efficient DNA repair through classical non-homologous end joining. These results indicate that PHF6 is a novel DNA damage response regulator that promotes end joining-mediated repair, thereby stimulating timely recovery from the G2 checkpoint.

| S-EPMC6944915 | biostudies-literature

Modeling damage complexity-dependent non-homologous end-joining repair pathway.

Project description:Non-homologous end joining (NHEJ) is the dominant DNA double strand break (DSB) repair pathway and involves several repair proteins such as Ku, DNA-PKcs, and XRCC4. It has been experimentally shown that the choice of NHEJ proteins is determined by the complexity of DSB. In this paper, we built a mathematical model, based on published data, to study how NHEJ depends on the damage complexity. Under an appropriate set of parameters obtained by minimization technique, we can simulate the kinetics of foci track formation in fluorescently tagged mammalian cells, Ku80-EGFP and DNA-PKcs-YFP for simple and complex DSB repair, respectively, in good agreement with the published experimental data, supporting the notion that simple DSB undergo fast repair in a Ku-dependent, DNA-PKcs-independent manner, while complex DSB repair requires additional DNA-PKcs for end processing, resulting in its slow repair, additionally resulting in slower release rate of Ku and the joining rate of complex DNA ends. Based on the numerous experimental descriptions, we investigated several models to describe the kinetics for complex DSB repair. An important prediction of our model is that the rejoining of complex DSBs is through a process of synapsis formation, similar to a second order reaction between ends, rather than first order break filling/joining. The synapsis formation (SF) model allows for diffusion of ends before the synapsis formation, which is precluded in the first order model by the rapid coupling of ends. Therefore, the SF model also predicts the higher number of chromosomal aberrations observed with high linear energy transfer (LET) radiation due to the higher proportion of complex DSBs compared to low LET radiation, and an increased probability of misrejoin following diffusion before the synapsis is formed, while the first order model does not provide a mechanism for the increased effectiveness in chromosomal aberrations observed.

| S-EPMC3919704 | biostudies-literature

TDP1 promotes assembly of non-homologous end joining protein complexes on DNA.

Project description:The repair of DNA double-strand breaks (DSB) is central to the maintenance of genomic integrity. In tumor cells, the ability to repair DSBs predicts response to radiation and many cytotoxic anti-cancer drugs. DSB repair pathways include homologous recombination and non-homologous end joining (NHEJ). NHEJ is a template-independent mechanism, yet many NHEJ repair products carry limited genetic changes, which suggests that NHEJ includes mechanisms to minimize error. Proteins required for mammalian NHEJ include Ku70/80, the DNA-dependent protein kinase (DNA-PKcs), XLF/Cernunnos and the XRCC4:DNA ligase IV complex. NHEJ also utilizes accessory proteins that include DNA polymerases, nucleases, and other end-processing factors. In yeast, mutations of tyrosyl-DNA phosphodiesterase (TDP1) reduced NHEJ fidelity. TDP1 plays an important role in repair of topoisomerase-mediated DNA damage and 3'-blocking DNA lesions, and mutation of the human TDP1 gene results in an inherited human neuropathy termed SCAN1. We found that human TDP1 stimulated DNA binding by XLF and physically interacted with XLF to form TDP1:XLF:DNA complexes. TDP1:XLF interactions preferentially stimulated TDP1 activity on dsDNA as compared to ssDNA. TDP1 also promoted DNA binding by Ku70/80 and stimulated DNA-PK activity. Because Ku70/80 and XLF are the first factors recruited to the DSB at the onset of NHEJ, our data suggest a role for TDP1 during the early stages of mammalian NHEJ.

| S-EPMC4673895 | biostudies-literature

Antagonistic relationship of NuA4 with the non-homologous end-joining machinery at DNA damage sites.

Project description:The NuA4 histone acetyltransferase complex, apart from its known role in gene regulation, has also been directly implicated in the repair of DNA double-strand breaks (DSBs), favoring homologous recombination (HR) in S/G2 during the cell cycle. Here, we investigate the antagonistic relationship of NuA4 with non-homologous end joining (NHEJ) factors. We show that budding yeast Rad9, the 53BP1 ortholog, can inhibit NuA4 acetyltransferase activity when bound to chromatin in vitro. While we previously reported that NuA4 is recruited at DSBs during the S/G2 phase, we can also detect its recruitment in G1 when genes for Rad9 and NHEJ factors Yku80 and Nej1 are mutated. This is accompanied with the binding of single-strand DNA binding protein RPA and Rad52, indicating DNA end resection in G1 as well as recruitment of the HR machinery. This NuA4 recruitment to DSBs in G1 depends on Mre11-Rad50-Xrs2 (MRX) and Lcd1/Ddc2 and is linked to the hyper-resection phenotype of NHEJ mutants. It also implicates NuA4 in the resection-based single-strand annealing (SSA) repair pathway along Rad52. Interestingly, we identified two novel non-histone acetylation targets of NuA4, Nej1 and Yku80. Acetyl-mimicking mutant of Nej1 inhibits repair of DNA breaks by NHEJ, decreases its interaction with other core NHEJ factors such as Yku80 and Lif1 and favors end resection. Altogether, these results establish a strong reciprocal antagonistic regulatory function of NuA4 and NHEJ factors in repair pathway choice and suggests a role of NuA4 in alternative repair mechanisms in situations where some DNA-end resection can occur in G1.

| S-EPMC8483352 | biostudies-literature

Nuclear TRADD prevents DNA damage-mediated death by facilitating non-homologous end-joining repair.

Project description:TNF receptor-associated death domain (TRADD) is an essential mediator of TNF receptor signaling, and serves as an adaptor to recruit other effectors. TRADD has been shown to cycle between the cytoplasm and nucleus due to its nuclear localization (NLS) and export sequences (NES). However, the underlying function of nuclear TRADD is poorly understood. Here we demonstrate that cytoplasmic TRADD translocates to DNA double-strand break sites (DSBs) during the DNA damage response (DDR). Deficiency of TRADD or its sequestration in cytosol leads to accumulation of ?H2AX-positive foci in response to DNA damage, which is reversed by nuclear TRADD expression. TRADD facilitates non-homologous end-joining (NHEJ) by recruiting NHEJ repair factors 53BP1 and Ku70/80 complex, whereas TRADD is dispensable for homologous recombination (HR) repair. Finally, an impaired nuclear localization of TRADD triggers cell death through the persistent activation of JNK and accumulation of reactive oxygen species (ROS). Thus, our findings suggest that translocation of TRADD to DSBs into the nucleus contributes to cell survival in response to DNA damage through an activation of DNA damage repair.

| S-EPMC5469829 | biostudies-other

Functional Roles of Homologous Recombination and Non-Homologous End Joining in DNA Damage Response and Microevolution in Cryptococcus neoformans.

Project description:DNA double-strand breaks (DSBs) are the most deleterious type of DNA lesions because they cause loss of genetic information if not properly repaired. In eukaryotes, homologous recombination (HR) and non-homologous end joining (NHEJ) are required for DSB repair. However, the relationship of HR and NHEJ in DNA damage stress is unknown in the radiation-resistant fungus Cryptococcus neoformans. In this study, we found that the expression levels of HR- and NHEJ-related genes were highly induced in a Rad53-Bdr1 pathway-dependent manner under genotoxic stress. Deletion of RAD51, which is one of the main components in the HR, resulted in growth under diverse types of DNA damage stress, whereas perturbations of KU70 and KU80, which belong to the NHEJ system, did not affect the genotoxic stresses except when bleomycin was used for treatment. Furthermore, deletion of both RAD51 and KU70/80 renders cells susceptible to oxidative stress. Notably, we found that deletion of RAD51 induced a hypermutator phenotype in the fluctuation assay. In contrast to the fluctuation assay, perturbation of KU70 or KU80 induced rapid microevolution similar to that induced by the deletion of RAD51. Collectively, Rad51-mediated HR and Ku70/Ku80-mediated NHEJ regulate the DNA damage response and maintain genome stability.

| S-EPMC8307084 | biostudies-literature

DROSHA is recruited to DNA damage sites by the MRN complex to promote non-homologous end joining.

Project description:The DNA damage response (DDR) is the signaling cascade that recognizes DNA double-strand breaks (DSBs) and promotes their resolution via the DNA repair pathways of non-homologous end joining (NHEJ) or homologous recombination (HR). We and others have shown that DDR activation requires DROSHA; however, whether DROSHA exerts its functions by associating with damage sites, what controls its recruitment, and how DROSHA influences DNA repair remains poorly understood. Here, we show that DROSHA associates with DSBs independently of transcription. Neither H2AX, nor ATM or DNA-PK kinase activities are required for recruitment of DROSHA to break sites. Rather, DROSHA interacts with RAD50, and inhibition of the MRN complex by mirin treatment abolishes this interaction. MRN complex inactivation by RAD50 knockdown or mirin treatment prevents DROSHA recruitment to DSBs and, as a consequence, also prevents 53BP1 (also known as TP53BP1) recruitment. During DNA repair, DROSHA inactivation reduces NHEJ and boosts HR frequency. Indeed, DROSHA knockdown also increases the association of downstream HR factors such as RAD51 to DNA ends. Overall, our results demonstrate that DROSHA is recruited at DSBs by the MRN complex and directs DNA repair towards NHEJ.

| S-EPMC8015226 | 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