RPRM negatively regulates ATM levels involving its phosphorylation mediated by CDK4/CDK6

Sensitizing cancer cells to radio- and chemotherapy remains a hot topic in cancer treatment. Here it is identified that Protein Reprimo (RPRM) negatively regulates the levels of ataxia-telangiectasia mutated (ATM) protein kinase, a master regulator of DNA damage response (DDR) in the presence of DNA double-strand breaks (DSBs), resulting in impaired DNA repair efficiency and enhanced cellular sensitivity to genotoxic agents. Mechanistically, although RPRM is primarily located in cytoplasm, it rapidly translocates to nucleus shortly after induced by X-irradiation, interacts with ATM and promotes the nuclear export and proteasomal degradation of ATM. The nuclear translocation of RPRM is associated with its phosphorylation at serine 98, which is mediated by cyclin-dependent kinases 4/6 (CDK4/6). Inhibition of CDK4/6 stabilizes RPRM and promotes its nuclear import, in turn enhances the nuclear export of ATM and the reduction of ATM levels. As a result, RPRM overexpression and its phosphorylation inhibition sensitize cells to genotoxic agents. Moreover, RPRM deficiency significantly increases resistance to radiation-induced damage both in vitro and in vivo. These findings establish a crucial regulatory mechanism in which ATM is negatively modulated by RPRM, suggesting that RPRM may serve as a novel target for both cancer therapy and radiation protection.

Cell Death and Survival Pathways Involving ATM Protein Kinase

Cell death is the ultimate form of cellular dysfunction, and is induced by a wide range of stresses including genotoxic stresses. During genotoxic stress, two opposite cellular reactions, cellular protection through DNA repair and elimination of damaged cells by the induction of cell death, can occur in both separate and simultaneous manners. ATM (ataxia telangiectasia mutated) kinase (hereafter referred to as ATM) is a protein kinase that plays central roles in the induction of cell death during genotoxic stresses. It has long been considered that ATM mediates DNA damage-induced cell death through inducing apoptosis. However, recent research progress in cell death modality is now revealing ATM-dependent cell death pathways that consist of not only apoptosis but also necroptosis, ferroptosis, and dysfunction of autophagy, a cellular survival mechanism. In this short review, we intend to provide a brief outline of cell death mechanisms in which ATM is involved, with emphasis on pathways other than apoptosis.

DNA Double-Strand Breaks Induced in Human Cells by Twelve Metallic Species: Quantitative Inter-Comparisons and Influence of the ATM Protein

Despite a considerable amount of data, the molecular and cellular bases of the toxicity due to metal exposure remain unknown. Recent mechanistic models from radiobiology have emerged, pointing out that the radiation-induced nucleo-shuttling of the ATM protein (RIANS) initiates the recognition and the repair of DNA double-strand breaks (DSB) and the final response to genotoxic stress. In order to document the role of ATM-dependent DSB repair and signalling after metal exposure, we applied twelve different metal species representing nine elements (Al, Cu, Zn Ni, Pd, Cd, Pb, Cr, and Fe) to human skin, mammary, and brain cells. Our findings suggest that metals may directly or indirectly induce DSB at a rate that depends on the metal properties and concentration, and tissue type. At specific metal concentration ranges, the nucleo-shuttling of ATM can be delayed which impairs DSB recognition and repair and contributes to toxicity and carcinogenicity. Interestingly, as observed after low doses of ionizing radiation, some phenomena equivalent to the biological response observed at high metal concentrations may occur at lower concentrations. A general mechanistic model of the biological response to metal exposure based on the nucleo-shuttling of ATM is proposed to describe the metal-induced stress response and to define quantitative endpoints for toxicity and carcinogenicity.

Chromosomal Location of Genes Differentially Expressed in Tumor Cells Surviving High-Dose X-ray Irradiation: A Preliminary Study on Radio-Fragile Sites

Altered gene expression is a common feature of tumor cells after irradiation. Our previous study showed that this phenomenon is not only an acute response to cytotoxic stress, instead, it was persistently detected in tumor cells that survived 10 Gy irradiation (IR cells). The current understanding is that DNA double-strand breaks (DSBs) are recognized by the phosphorylation of histone H2AX (H2AX) and triggers the ataxia-telangiectasia mutated (ATM) protein or the ATM- and Rad3-related (ATR) pathway, which activate or inactivate the DNA repair or apoptotic or senescence related molecules and causes the expression of genes in many instances. However, because changes in gene expression persist after passaging in IR cells, it may be due to the different pathways from these transient intracellular signaling pathways caused by DSBs. We performed microarray analysis of 30,000 genes in radiation-surviving cells (H1299-IR and MCF7-IR) and found an interesting relation between altered genes and their chromosomal loci. These loci formed a cluster on the chromosome, especially on 1q21 and 6p21-p22 in both irradiated cell lines. These chromosome sites might be regarded as “radio-fragile” sites.

Oxidized ATM Protein Kinase Promotes the Proliferation of Breast CAF Through Upregulating the SCD1 Gene Expression by Activating Core Proliferation-Related Pathways

PurposeStearoyl-CoA desaturase 1 (SCD1) is a key rate-limiting enzyme involved in the biosynthesis of monounsaturated fatty acids, which mediates lipid metabolism and participates in the proliferation of cancer cells. As a redox sensor, ATM protein kinase is also involved in DNA repair and cell proliferation regulation. Previous studies have shown that oxidized ATM protein kinase can facilitates the proliferation of breast CAF via by regulating the cell glucose and protein metabolism. However, lipid metabolism is essential for cell proliferation. Whether the oxidized ATM protein kinase can regulate the expression of the SCD gene to promote the lipid metabolism is unknown.MethodsWe use immunohistochemical techniques to observe the level of SCD1 and oxidized ATM protein kinase, MTT Assay and flow cytometric analysis observes the proliferation of breast CAFs, and Western Blot techniques analyze the expression of SCD gene expression.ResultsThe expression levels of SCD1 protein expression in breast cancer were closely related to oxidized ATM expression (P < 0.05). And SCD promotes the proliferation of breast CAF cells. The oxidized ATM protein kinase can promote the proliferation of breast CAFs by upregulation of SCD protein expression levels.ConclusionOur results indicate that oxidized ATM protein kinase may play a central role in promoting the proliferation of breast CAF by coordinating glucose, protein and lipid metabolism. oxidized ATM can facilitate the breast CAFs Lipid metabolism by upregulating the SCD1 gene expression.

Detection of Genes in Arabidopsis thaliana L. Responding to DNA Damage from Radiation and Other Stressors in Spaceflight

Ionizing radiation present in extraterrestrial environment is an important factor that affects plants grown in spaceflight. Pearson correlation-based gene regulatory network inferencing from transcriptional responses of the plant Arabidopsis thaliana L. grown in real and simulated spaceflight conditions acquired by GeneLab, followed by topological and spectral analysis of the networks is performed. Gene regulatory subnetworks are extracted for DNA damage response processes. Analysis of radiation-induced ATR/ATM protein–protein interactions in Arabidopsis reveals interaction profile similarities under low radiation doses suggesting novel mechanisms of DNA damage response involving non-radiation-induced genes regulating other stress responses in spaceflight. The Jaccard similarity index shows that the genes AT2G31320, AT4G21070, AT2G46610, and AT3G27060 perform similar functions under low doses of radiation. The incremental association Markov blanket method reveals non-radiation-induced genes linking DNA damage response to root growth and plant development. Eighteen radiation-induced genes and sixteen non-radiation-induced gene players have been identified from the ATR/ATM protein interaction complexes involved in heat, salt, water, osmotic stress responses, and plant organogenesis. Network analysis and logistic regression ranking detected AT3G27060, AT1G07500, AT5G66140, and AT3G21280 as key gene players involved in DNA repair processes. High atomic weight, high energy, and gamma photon radiation result in higher intensity of DNA damage response in the plant resulting in elevated values for several network measures such as spectral gap and girth. Nineteen flavonoid and carotenoid pigment activations involved in pigment biosynthesis processes are identified in low radiation dose total light spaceflight environment but are not found to have significant regulations under very high radiation dose environment.

ATM controls the extent of DNA end resection by eliciting sequential posttranslational modifications of CtIP

DNA end resection is a critical step in the repair of DNA double-strand breaks (DSBs) via homologous recombination (HR). However, the mechanisms governing the extent of resection at DSB sites undergoing homology-directed repair remain unclear. Here, we show that, upon DSB induction, the key resection factor CtIP is modified by the ubiquitin-like protein SUMO at lysine 578 in a PIAS4-dependent manner. CtIP SUMOylation occurs on damaged chromatin and requires prior hyperphosphorylation by the ATM protein kinase. SUMO-modified hyperphosphorylated CtIP is targeted by the SUMO-dependent E3 ubiquitin ligase RNF4 for polyubiquitination and subsequent degradation. Consequently, disruption of CtIP SUMOylation results in aberrant accumulation of CtIP at DSBs, which, in turn, causes uncontrolled excessive resection, defective HR, and increased cellular sensitivity to DSB-inducing agents. These findings reveal a previously unidentified regulatory mechanism that regulates CtIP activity at DSBs and thus the extent of end resection via ATM-dependent sequential posttranslational modification of CtIP.