Search

Research has been conducted on the human glioma cell line extracts which lack O (super)6 -methylguanine DNA-methyltransferase. The ability of these extracts to execute strand incision at various base mismatches in model DNA has been investigated and the results are discussed.

Herein we used single-cell observation methods to gain insight into the roles of p16INK4A and p21WAF1 (hereafter p16 and p21) in replicative senescence and ionizing radiation-induced accelerated senescence in human [normal, ataxia telangiectasia (AT) and Li-Fraumeni syndrome (LFS)] fibroblast strains. Cultures of all strains entered a state of replicative senescence at late passages, as evident from inhibition of growth, acquisition of flattened and enlarged cell morphology, and positive staining for senescence-associated β-galactosidase. In addition, proliferating early-passage cultures of these strains exhibited accelerated senescence in response to ionizing radiation. Immunofluorescence microscopy revealed the heterogeneous expression of p16 in normal and AT fibroblast strains, with the majority of the cells exhibiting undetectable levels of p16 irrespective of in vitro culture age. Importantly, replicative senescence as well as accelerated senescence triggered by ionizing radiation were accompanied by sustained nuclear accumulation of p21, but did not correlate with p16 expression in p53-proficient (normal and AT) fibroblasts. In p53-deficient (LFS) fibroblasts, on the other hand, replicative senescence and ionizing radiation-triggered accelerated senescence strongly correlated with expression of p16 but not of p21. Furthermore, senescence in LFS fibroblasts was associated with genomic instability encompassing polyploidy. Our findings are compatible with a model in which p16 serves as a backup regulator of senescence, triggering this response preferentially in the absence of wild-type p53 activity. The possibility that one of the tumor-suppressor functions of p16 may be associated with genomic instability, preventing the emergence of malignant progeny from polyploid giant cells, is also supported by these results. J. Cell. Physiol. 223: 57–67, 2010. © 2009 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Ataxia telangiectasia (AT) is a human hereditary syndrome whose underlying gene product, ataxia telangiectasia mutated (ATM) protein kinase, is involved in multiple intracellular signaling pathways. We demonstrated previously that AT fibroblasts are defective in intracellular Ca2+ mobilization in response to both stress-inducing and mitogenic stimuli. To extend these findings, normal and AT cells were exposed to serum in the presence of different concentrations of extracellular Ca2+ ([Ca2+]o), and release of intracellular Ca2+, activation of calmodulin-dependent protein kinase II and phosphorylation of kinases ERK1 and 2 were monitored. When maintained in high [Ca2+]o (0.42 mM), normal fibroblasts responded to serum introduction more rapidly and efficiently than did AT cells. Unexpectedly, decreasing the [Ca2+]o in the medium had a diametrically opposite effect. Under low [Ca2+]o (0.0022 mM) conditions, normal cells were slow and inefficient in their responses, whereas AT cells showed a substantial improvement in all three end points. These findings demonstrate that loss of ATM kinase function deregulates the extracellular calcium-sensing receptor (CaR). This malfunction presumably arises from a post-transcriptional event, since CaR mRNA proved to be normal in AT cells. Together, our data suggest that ATM may mediate cell response to mitogenic factors by tightly regulating the set point of the CaR and thereby modulating the crosstalk between this metabotropic receptor and growth factor receptors. Alternatively, the faulty sensing of extracellular calcium in AT cells may be secondary to a state of chronic oxidative stress attributable to ATM deficiency.Oncogene (2003) 22, 471–475. doi:10.1038/sj.onc.1206167 [ABSTRACT FROM AUTHOR]