SIRT1 modulates cell cycle progression by regulating CHK2 acetylation-phosphorylation.

Both the stress-response protein, SIRT1, and the cell cycle checkpoint kinase, CHK2, play critical roles in aging and cancer via the modulation of cellular homeostasis and the maintenance of genomic integrity. However, the underlying mechanism linking the two pathways remains elusive. Here, we show that SIRT1 functions as a modifier of CHK2 in cell cycle control. Specifically, SIRT1 interacts with CHK2 and deacetylates it at lysine 520 residue, which suppresses CHK2 phosphorylation, dimerization, and thus activation. SIRT1 depletion induces CHK2 hyperactivation-mediated cell cycle arrest and subsequent cell death. In vivo, genetic deletion of Chk2 rescues the neonatal lethality of Sirt1-/- mice, consistent with the role of SIRT1 in preventing CHK2 hyperactivation. Together, these results suggest that CHK2 mediates the function of SIRT1 in cell cycle progression, and may provide new insights into modulating cellular homeostasis and maintaining genomic integrity in the prevention of aging and cancer.

Chk2 deficiency alleviates irradiation-induced taste dysfunction by inhibiting p53-dependent apoptosis.

OBJECTIVE: Taste dysfunction is one of the most common complications following radiotherapy, which leads to decreased appetite and life quality of patients suffering from head and neck cancer. The aim of this study was to investigate the role of checkpoint kinase 2 (Chk2) deficiency in irradiation-induced taste dysfunction. MATERIALS AND METHODS: Alterations in oxidative stress, DNA damage, and potential signaling pathway were compared between Chk2-deficient (Chk2-/- ) mice and their wild-type (WT) littermates pre-irradiation and 7 and 30 days postirradiation by biochemistry and immunohistochemistry. RESULTS: Chk2-/- mice showed less loss of type II and type III taste cells, lower expression of p53, caspase-3, and cleaved caspase-3, and lower apoptosis levels. However, no significant differences in H2 O2 and MDA concentrations, T-SOD and GSH-Px activities, and expression of SOD1, SOD2, and 8-OHdG were detected in the taste buds of Chk2-/- mice as compared to those of WT mice. CONCLUSION: Chk2 deficiency downregulated the expression of p53 and inhibited cellular apoptosis, partly contributing to the radioprotective effect on taste cells, but did not alter oxidative stress levels, antioxidant ability, and oxidative DNA damage in taste buds.

Associations between mutations of the cell cycle checkpoint kinase 2 gene and gastric carcinogenesis.

Gastric cancer is the most common malignant tumor of the digestive system. The etiology of gastric cancer is complex, and susceptibility at the genetic level remains to be fully elucidated in genetic investigations. In the present study, mutations of the cell cycle checkpoint kinase 2 (CHEK2) gene and its association with gastric cancer were examined. Reverse transcription­quantitative polymerase chain reaction technology was used to detect the expression of CHEK2 and it was found that the expression of CHEK2 was low in gastric cancer. Using sequencing analysis, it was found that the low expression level of CHEK2 was associated with expression of its mutation. The present study also established a CHEK2­overexpressing mutant and confirmed that CHEK2 promoted gastric cancer cell proliferation. Overexpression of the CHEK2 mutation was confirmed to promote cancer cell migration and invasion. Furthermore, western blot analysis results revealed that overexpression of the CHEK2 mutation downregulated E­cadherin and upregulated vimentin expression, indicating the mechanism underlying the altered biological behavior. These results suggested that there was a correlation between mutation of the CHEK2 gene and gastric cancer, and provided an experimental basis for antitumor drug investigation and development according to its mutation target.

Checkpoint kinase 2 is required for efficient immunoglobulin diversification.

Maintenance of genome integrity relies on multiple DNA repair pathways as well as on checkpoint regulation. Activation of the checkpoint kinases Chk1 and Chk2 by DNA damage triggers cell cycle arrest and improved DNA repair, or apoptosis in case of excessive damage. Chk1 and Chk2 have been reported to act in a complementary or redundant fashion, depending on the physiological context. During secondary immunoglobulin (Ig) diversification in B lymphocytes, DNA damage is abundantly introduced by activation-induced cytidine deaminase (AID) and processed to mutations in a locus-specific manner by several error-prone DNA repair pathways. We have previously shown that Chk1 negatively regulates Ig somatic hypermutation by promoting error-free homologous recombination and Ig gene conversion. We now report that Chk2 shows opposite effects to Chk1 in the regulation of these processes. Chk2 inactivation in B cells leads to decreased Ig hypermutation and Ig class switching, and increased Ig gene conversion activity. This is linked to defects in non-homologous end joining and increased Chk1 activation upon interference with Chk2 function. Intriguingly, in the context of physiological introduction of substantial DNA damage into the genome during Ig diversification, the 2 checkpoint kinases thus function in an opposing manner, rather than redundantly or cooperatively.

Chk2 prevents mitotic exit when the majority of kinetochores are unattached.

The spindle checkpoint delays exit from mitosis in cells with spindle defects. In this paper, we show that Chk2 is required to delay anaphase onset when microtubules are completely depolymerized but not in the presence of relatively few unattached kinetochores. Mitotic exit in Chk2-deficient cells correlates with reduced levels of Mps1 protein and increased Cdk1-tyrosine 15 inhibitory phosphorylation. Chk2 localizes to kinetochores and is also required for Aurora B-serine 331 phosphorylation in nocodazole or unperturbed early prometaphase. Serine 331 phosphorylation contributed to prometaphase accumulation in nocodazole after partial Mps1 inhibition and was required for spindle checkpoint establishment at the beginning of mitosis. In addition, expression of a phosphomimetic S331E mutant Aurora B rescued chromosome alignment or segregation in Chk2-deficient cells. We propose that Chk2 stabilizes Mps1 and phosphorylates Aurora B-serine 331 to prevent mitotic exit when most kinetochores are unattached. These results highlight mechanisms of an essential function of Chk2 in mitosis.

Role of DNA damage response pathways in preventing carcinogenesis caused by intrinsic replication stress.

Defective DNA replication can result in genomic instability, cancer and developmental defects. To understand the roles of DNA damage response (DDR) genes on carcinogenesis in mutants defective for core DNA replication components, we utilized the Mcm4(Chaos3/Chaos3) ('Chaos3') mouse model that, by virtue of an amino-acid alteration in MCM4 that destabilizes the MCM2-7 DNA replicative helicase, has fewer dormant replication origins and an increased number of stalled replication forks. This leads to genomic instability and cancer in most Chaos3 mice. We found that animals doubly mutant for Chaos3 and components of the ataxia telangiectasia-mutated (ATM) double-strand break response pathway (Atm, p21/Cdkn1a and Chk2/Chek2) had decreased tumor latency and/or increased tumor susceptibility. Tumor latency and susceptibility differed between genetic backgrounds and genders, with females demonstrating an overall greater cancer susceptibility to Atm and p21 deficiency than males. Atm deficiency was semilethal in the Chaos3 background and impaired embryonic fibroblast proliferation, suggesting that ATM drug inhibitors might be useful against tumors with DNA replication defects. Hypomorphism for the 9-1-1 component Hus1 did not affect tumor latency or susceptibility in Chaos3 animals, and tumors in these mice did not exhibit impaired ATR pathway signaling. These and other data indicate that under conditions of systemic replication stress, the ATM pathway is particularly important both for cancer suppression and viability during development.