R-loops and impaired autophagy trigger cGAS-dependent inflammation via micronuclei formation in Senataxin-deficient cells.

Senataxin is an evolutionarily conserved DNA/RNA helicase, whose dysfunctions are linked to neurodegeneration and cancer. A main activity of this protein is the removal of R-loops, which are nucleic acid structures capable to promote DNA damage and replication stress. Here we found that Senataxin deficiency causes the release of damaged DNA into extranuclear bodies, called micronuclei, triggering the massive recruitment of cGAS, the apical sensor of the innate immunity pathway, and the downstream stimulation of interferon genes.

Sex specific regulation of TSPY-Like 2 in the DNA damage response of cancer cells.

Females have a lower probability to develop somatic cancers and a better response to chemotherapy than males. However, the reasons for these differences are still not well understood. The X-linked gene TSPY-Like 2 (TSPYL2) encodes for a putative tumor suppressor protein involved in cell cycle regulation and DNA damage response (DDR) pathways.

Sex disparities in DNA damage response pathways: Novel determinants in cancer formation and therapy.

Cancer incidence and survival are different between men and women. Indeed, females have a lesser risk and a better prognosis than males in many tumors unrelated to reproductive functions. Although the reasons for these disparities are still unknown, they constitute an important starting point for the development of personalized cancer therapies.

Targeting SMYD3 to Sensitize Homologous Recombination-Proficient Tumors to PARP-Mediated Synthetic Lethality.

is frequently overexpressed in a wide variety of cancers. Indeed, its inactivation reduces tumor growth in preclinical animal models. However, extensive characterization failed to clarify function in cancer cells, although confirming its importance in carcinogenesis.

TSPYL2 is a novel regulator of SIRT1 and p300 activity in response to DNA damage.

Protein acetylation and deacetylation events are finely regulated by lysine-acetyl-transferases and lysine-deacetylases and constitute an important tool for the activation or inhibition of specific cellular pathways. One of the most important lysine-acetyl-transferases is p300, which is involved in the regulation of gene expression, cell growth, DNA repair, differentiation, apoptosis, and tumorigenesis. A well-known target of p300 is constituted by the tumor suppressor protein p53, which plays a critical role in the maintenance of genomic stability and whose activity is known to be controlled by post-translational modifications, among which acetylation.

Cell cycle and apoptosis regulator 2 at the interface between DNA damage response and cell physiology.

Cell cycle and apoptosis regulator 2 (CCAR2 or DBC1) is a human protein recently emerged as a novel and important player of the DNA damage response (DDR). Indeed, upon genotoxic stress, CCAR2, phosphorylated by the apical DDR kinases ATM and ATR, increases its binding to the NAD+-dependent histone deacetylase SIRT1 and inhibits SIRT1 activity. This event promotes the acetylation and activation of p53, a SIRT1 target, and the subsequent induction of p53 dependent apoptosis.

A novel crosstalk between CCAR2 and AKT pathway in the regulation of cancer cell proliferation.

Human CCAR2 has recently emerged as having a pivotal role in the DNA damage response, promoting apoptosis and repair of heterochromatic DNA breaks. However, less is known about the function of CCAR2 in tumor formation and cancer progression. Here, we demonstrate, for the first time, that CCAR2 loss inhibits the proliferation of cancer cells, but preserves the growth of normal cells.

Retina-derived POU domain factor 1 coordinates expression of genes relevant to renal and neuronal development.

Retina-derived POU domain Factor 1 (RPF-1), a member of POU transcription factor family, is encoded by POU6F2 gene, addressed by interstitial deletions at chromosome 7p14 in Wilms tumor (WT). Its expression has been detected in developing kidney and nervous system, suggesting an early role for this gene in regulating development of these organs. To investigate into its functions and determine its role in transcriptional regulation, we generated an inducible stable transfectant from HEK293 cells.

CCAR2/DBC1 is required for Chk2-dependent KAP1 phosphorylation and repair of DNA damage.

Cell cycle and apoptosis regulator 2 (CCAR2, formerly known as DBC1) is a nuclear protein largely involved in DNA damage response, apoptosis, metabolism, chromatin structure and transcription regulation. Upon DNA lesions, CCAR2 is phosphorylated by the apical kinases ATM/ATR and this phosphorylation enhances CCAR2 binding to SIRT1, leading to SIRT1 inhibition, p53 acetylation and p53-dependent apoptosis. Recently, we found that also the checkpoint kinase Chk2 and the proteasome activator REGγ are required for efficient CCAR2-mediated inhibition of SIRT1 and induction of p53-dependent apoptosis.

CHK2 kinase in the DNA damage response and beyond.

The serine/threonine kinase CHK2 is a key component of the DNA damage response. In human cells, following genotoxic stress, CHK2 is activated and phosphorylates >20 proteins to induce the appropriate cellular response, which, depending on the extent of damage, the cell type, and other factors, could be cell cycle checkpoint activation, induction of apoptosis or senescence, DNA repair, or tolerance of the damage. Recently, CHK2 has also been found to have cellular functions independent of the presence of nuclear DNA lesions.

Chk2 and REGγ-dependent DBC1 regulation in DNA damage induced apoptosis.

Human DBC1 (Deleted in Breast Cancer 1; KIAA1967; CCAR2) is a protein implicated in the regulation of apoptosis, transcription and histone modifications. Upon DNA damage, DBC1 is phosphorylated by ATM/ATR on Thr454 and this modification increases its inhibitory interaction with SIRT1, leading to p53 acetylation and p53-dependent apoptosis. Here, we report that the inhibition of SIRT1 by DBC1 in the DNA damage response (DDR) also depends on Chk2, the transducer kinase that is activated by ATM upon DNA lesions and contributes to the spreading of DNA damage signal.

DBC1 phosphorylation by ATM/ATR inhibits SIRT1 deacetylase in response to DNA damage.

Human DBC1 (deleted in breast cancer-1; KIAA1967) is a nuclear protein that, in response to DNA damage, competitively inhibits the NAD(+)-dependent deacetylase SIRT1, a regulator of p53 apoptotic functions in response to genotoxic stress. DBC1 depletion in human cells increases SIRT1 activity, resulting in the deacetylation of p53 and protection from apoptosis. However, the mechanisms regulating this process have not yet been determined.

A protein phosphatase feedback mechanism regulates the basal phosphorylation of Chk2 kinase in the absence of DNA damage.

The checkpoint kinase Chk2 is an effector component of the ATM-dependent DNA damage response (DDR) pathway. The activation of Chk2 by genotoxic stress involves its phosphorylation on T68 by ATM and additional auto/transphosphorylations. Here we demonstrate that in unperturbed cells, chemical inhibition of Chk2 by VRX0466617 (VRX) enhances the phosphorylation of Chk2-T68 throughout the cell cycle phases.

REGgamma/PA28gamma proteasome activator interacts with PML and Chk2 and affects PML nuclear bodies number.

REGgamma (also called PA28gamma or PSME3) is a proteasome activator involved in the degradation of several proteins that regulate cell cycle and transcription. Recently, we demonstrated that this protein has a role also in the maintenance of chromosomal stability and in the response to spindle damaging agents. Here we report for the first time that REGgamma interacts with the promyelocytic leukemia protein (PML), accumulates in PML nuclear bodies (PML-NBs), but it does not play any role in normal or arsenic-induced PML degradation.

The shelterin protein TRF2 inhibits Chk2 activity at telomeres in the absence of DNA damage.

The shelterin complex [1] shapes and protects telomeric DNA from being processed as double strand breaks (DSBs) [2, 3]. Here we show that in human undamaged cells, a fraction of the kinase Chk2, a downstream target of ATM and mediator of checkpoint responses and senescence [4, 5], physically interacts with the shelterin subunit TRF2 and colocalizes with this complex at chromosome ends. This interaction, enhanced by TRF2 binding to telomeric DNA, inhibits the activation and senescence-induced function of Chk2 by a mechanism in which TRF2 binding to the N terminus of Chk2 surrounding Thr68 hinders the phosphorylation of this priming site.

REGgamma proteasome activator is involved in the maintenance of chromosomal stability.

REGgamma is a member of the 11S regulatory particle that activates the 20S proteasome. Studies in REGgamma deficient mice indicated an additional role for this protein in cell cycle regulation and proliferation control. In this paper we demonstrate that REGgamma protein is equally expressed throughout the cell cycle, but undergoes a distinctive subcellular localization at mitosis.

Biochemical and cellular characterization of VRX0466617, a novel and selective inhibitor for the checkpoint kinase Chk2.

VRX0466617 is a novel selective small-molecule inhibitor for Chk2 discovered through a protein kinase screening program. In this study, we provide a detailed biochemical and cellular characterization of VRX0466617. We show that VRX0466617 blocks the enzymatic activity of recombinant Chk2, as well as the ionizing radiation (IR)-induced activation of Chk2 from cells pretreated with the compound, at doses between 5 and 10 micromol/L.

Differential roles of ATM- and Chk2-mediated phosphorylations of Hdmx in response to DNA damage.

The p53 tumor suppressor plays a major role in maintaining genomic stability. Its activation and stabilization in response to double strand breaks (DSBs) in DNA are regulated primarily by the ATM protein kinase. ATM mediates several posttranslational modifications on p53 itself, as well as phosphorylation of p53's essential inhibitors, Hdm2 and Hdmx.

DNA damage-induced cell cycle regulation and function of novel Chk2 phosphoresidues.

Chk2 kinase is activated by DNA damage to regulate cell cycle arrest, DNA repair, and apoptosis. Phosphorylation of Chk2 in vivo by ataxia telangiectasia-mutated (ATM) on threonine 68 (T68) initiates a phosphorylation cascade that promotes the full activity of Chk2. We identified three serine residues (S19, S33, and S35) on Chk2 that became phosphorylated in vivo rapidly and exclusively in response to ionizing radiation (IR)-induced DNA double-strand breaks in an ATM- and Nbs1-dependent but ataxia telangiectasia- and Rad3-related-independent manner.

Activation of ATM and Chk2 kinases in relation to the amount of DNA strand breaks.

The diverse checkpoint responses to DNA damage may reflect differential sensitivities by molecular components of the damage-signalling network to the type and amount of lesions. Here, we determined the kinetics of activation of the checkpoint kinases ATM and Chk2 (the latter substrate of ATM) in relation to the initial yield of genomic DNA single-strand (SSBs) and double-strand breaks (DSBs). We show that doses of gamma-radiation (IR) as low as 0.

MRE11 mutations and impaired ATM-dependent responses in an Italian family with ataxia-telangiectasia-like disorder.

Hypomorphic mutations of the MRE11 gene are the hallmark of the radiosensitive ataxia-telangiectasia-like disorder (ATLD). Here, we describe a new family with two affected siblings, ATLD5 and ATLD6, now aged 37 and 36, respectively. They presented with late onset cerebellar degeneration slowly progressing until puberty and absence of telangiectasias, and were cancer-free.

Karyopherin-alpha2 protein interacts with Chk2 and contributes to its nuclear import.

Chk2 is a nuclear protein kinase involved in the DNA damage-induced ataxia telangiectasia mutated-dependent checkpoint arrest at multiple cell cycle phases. Searching for Chk2-binding proteins by a yeast two-hybrid system, we identified a strong interaction with karyopherin-alpha2 (KPNA-2), a gene product involved in active nuclear import of proteins bearing a nuclear localization signal (NLS). This finding was confirmed by glutathione S-transferase pull-down and co-immunoprecipitation assays.

The cell cycle-regulated protein human GTSE-1 controls DNA damage-induced apoptosis by affecting p53 function.

GTSE-1 (G2 and S phase-expressed-1) protein is specifically expressed during S and G2 phases of the cell cycle. It is mainly localized to the microtubules and when overexpressed delays the G2 to M transition. Here we report that human GTSE-1 (hGTSE-1) protein can negatively regulate p53 transactivation function, protein levels, and p53-dependent apoptosis.