Cdk1 restrains NHEJ through phosphorylation of XRCC4-like factor Xlf1

Eukaryotic cells use two principal mechanisms for repairing DNA double-strand breaks (DSBs): homologous recombination (HR) and nonhomologous end-joining (NHEJ). DSB repair pathway choice is strongly regulated during the cell cycle. Cyclin-dependent kinase 1 (Cdk1) activates HR by phosphorylation of key recombination factors. However, a mechanism for regulating the NHEJ pathway has not been established. Here, we report that Xlf1, a fission yeast XLF ortholog, is a key regulator of NHEJ activity in the cell cycle. We show that Cdk1 phosphorylates residues in the C terminus of Xlf1 over the course of the cell cycle. Mutation of these residues leads to the loss of Cdk1 phosphorylation, resulting in elevated levels of NHEJ repair in vivo. Together, these data establish that Xlf1 phosphorylation by Cdc2Cdk1 provides a molecular mechanism for downregulation of NHEJ in fission yeast and indicates that XLF is a key regulator of end-joining processes in eukaryotic organisms

Plant-derived tetranortriterpenoid, methyl angolensate activates apoptosis and prevents ehrlich ascites carcinoma induced tumorigenesis in mice

Background: Cancer is a leading health problem throughout the world. For decades, natural plant products have been playing promising roles as anticancer agents. Objective: The present study aims to investigate the chemotherapeutic potential of Methyl Angolensate (MA), purified from Soymida febrifuga in mice bearing carcinoma and examines the molecular basis for its anticancer actions. Study Design: The inhibitory effects of MA treatment on the survival of mice bearing Carcinoma and adverse side effects of MA treatment in mice were analyzed. Methods: Tumor volume, life span, histopathology, Immunohistochemical (IHC) analysis, estimation of liver enzyme, alkaline phosphatase and metabolites, creatinine and urea. Results: Oral administration of MA in mice with Ehrlich Ascites Carcinoma showed significant inhibition of tumor growth compared to untreated mice. We observed a significant increase in the life span (∼4-fold) of tumor bearing animals following treatment with MA. MA affected tumor cell proliferation by activating intrinsic pathway of apoptosis without imparting any side effect on normal cells. MA treatment in mice showed no major side effects. Conclusion: MA treatment showed significant inhibition of tumor growth by inducing apoptosis as well increased life span of mice, with no adverse side effects to normal cells. Altogether, the present in vivo study provides new insights of MA serving as a cancer chemotherapeutic agent

PARP1 promoter links cell cycle progression with adaptation to oxidative environment

Although electrophiles are considered as detrimental to cells, accumulating recent evidence indicates that proliferating non-cancerous and particularly cancerous cells utilize these agents for pro-survival and cell cycle promoting signaling. Hence, the redox shift to mild oxidant release must be balanced by multiple defense mechanisms. Our latest findings demonstrate that cell cycle progression, which dictates oxidant level in stress-free conditions, determines PARP1 transcription. Growth modulating factors regulate CDK4/6-RBs-E2Fs axis. In cells arrested in G1 and G0, RB1-E2F1 and RBL2-E2F4 dimers recruit chromatin remodelers such as HDAC1, SWI/SNF and PRC2 to condense chromatin and turn off transcription. Release of retinoblastoma-based repressive complexes from E2F-dependent gene promoters in response to cell transition to S phase enables transcription of PARP1. This enzyme contributes to repair of oxidative DNA damage by supporting several strand break repair pathways and nucleotide or base excision repair pathways, as well as acting as a co-activator of transcription factors such as NRF2 and HIF1a, which control expression of antioxidant enzymes involved in removal of electrophiles and secondary metabolites. Furthermore, PARP1 is indispensible for transcription of the pro-survival kinases MAP2K6, ERK1/2 and AKT1, and for maintaining MAPK activity by suppressing transcription of the MAPK inhibitor, MPK1. In summary, cell cycle controlled PARP1 transcription helps cells to adapt to a pro-oxidant redox shift

Identification of a novel BCL2-specific inhibitor that binds predominantly to the BH1 domain

The antiapoptotic protein BCL 2 is overexpressed in several cancers and contributes to prolonged cell survival and chemoresistance, lending itself as an excellent target for cancer therapy. Here, we report the design, synthesis, and characterization of Disarib, a novel BCL 2 inhibitor. Disarib showed selective cytotoxicity in BCL 2 high cancer cell lines, and CLL patient primary cells, as compared to BCL 2 low cell lines. BCL 2 knockdown in cells rendered remarkable resistance to Disarib, while sensitivity was regained upon its ectopic expression, establishing target specificity. In silico , biochemical and biophysical studies demonstrated strong affinity of Disarib to BCL 2, but not to other antiapoptotic BCL 2 family members viz., BCL ‐xL , BCL 2A1 etc. Interestingly, biophysical studies showed that BH 1 domain deletion mutant demonstrated ~ 67‐fold reduction in BCL 2‐Disarib interaction, while it was only ~ 20‐fold in the case of BH 3 deletion mutant, suggesting predominant involvement of the BH 1 domain for Disarib binding. Thus, we report identification of a novel BCL 2 inhibitor with a unique mechanism of BCL 2 inhibition, as opposed to the well‐studied BH 3 domain targeting

A natural compound, methyl angolensate, induces mitochondrial pathway of apoptosis in Daudi cells

Natural products discovered from medicinal plants have played an important role in the treatment of cancer. In an effort to identify novel small molecules which can affect the proliferation of lymphoma cells, we tested Methyl Angolensate (MA), a plant derived tetranortriterpenoid, purified from the crude extract of the root callus of Soymida febrifuga commonly known as Indian red wood tree. We have tested MA for its cytotoxic properties on Burkitt’s lymphoma cell lines, using various cellular assays. We observed that MA induces cytotoxicity in Daudi cells in a dose-dependent manner using trypan blue, MTT and LDH assays. We find that the treatment with MA led to activation of DNA double-strand break repair proteins including KU70 and KU80, suggesting the activation of nonhomologous DNA end joining pathway in surviving cells. Further, we find that methyl angolensate could induce apoptosis by cell cycle analysis, annexin V-FITC staining, DNA fragmentation and PARP cleavage. Besides, MA treatment led to reactive oxygen species generation and loss of mitochondrial transmembrane potential. These results suggest the activation of mitochondrial pathway of apoptosis. Hence, we identify MA as a potential chemotherapeutic agent against Daudi cells