p53 Tumor Suppressor Pathway

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The eukaryote genome constantly faces the threat of damage from exogenous and endogenous mutagens. Mammalian cells, therefore, have evolved an intricate network of defenses to maintain genomic stability. p53 is at the crossroads of these defense pathways. Protection of Telomeres 1(POT1)is a single stranded DNA binding component of the shelterin complex, which is essential to the formation of a telomere-specific t-loop structure and functions as a regulator of telomere length. Consistent with observations that Telomere-Related Factor-2 (TRF2) also functions in these processes, we reported that POT1 and TRF2 cooperated to protect telomeres and regulate cellular senescence and apoptosis. The siRNA knockdown of POT1 resulted in the loss of telomeric single-stranded overhangs (3' overhangs), chromosomal instability, apoptosis in breast cancer cells, and cellular senescence in normal human fibroblasts. POT1 physically interacted with TRF2 and protected against the dominant-negative TRF2-induced telomere dysfunction. We also found that a C-terminally truncated variant of human POT1 (v5) protected telomeres and prevented cellular senescence as effeciently as the full-length POT1, but in mechanistically different manners: (a) the full-length POT1, but not v5, functions through the maintenance of 3' overhangs; (b) p53 is dispensible to v5 knockdown-induced senescence; and (c) v5 functions at only a fraction of telomeres. Based on these results, we propose that human telomeres are cooperatively regulated by the functionally distinct POT1 variants. The telomere-protective variant v5 is preferentially expressed in mismatch repair-deficient tumors, which have stable chromosomes, suggesting the role of v5 in chromosome stability in human cancers. We are continuing to study p53-mediated replicative senescence modulated by POT1 or p53 isoforms that govern telomere attrition. The finite proliferative potential of normal human cells leads to replicative cellular senescence, which is a critical barrier in tumour progression in vivo. We have shown that human p53 isoforms (delta 133p53 and p53beta) function in an endogenous regulatory mechanism for p53-mediated replicative senescence. Induced p53beta and diminished delta133p53 were associated with replicative senescence, but not oncogene-induced senescence, in normal human fibroblasts. The replicatively senescent fibroblasts also expressed increased levels of miR-34a, a p53-induced microRNA, the antisense inhibition of which delayed the onset of replicative senescence. The short interfering RNA (siRNA)- mediated knockdown of endogenous delta133p53 induced cellular senescence, which was attributed to the regulation of p21waf1 and other p53 transcriptional target genes. In overexpression experiments, whereas p53beta cooperated with full-length p53 to accelarate cellular senescence, delta133p53 repressed miR-34a expression and extended the cellular replicative lifespan, providing a functional connection of this microRNA to the p53 isoform-mediated regulation of senescence. The senescence-associated signature of p53 isoform expression (that is, elevated p53beta and reduced delta133p53) was observed in vivo in colon adenomas with senescent phenotypes. The increased delta13p53 and decreased p53beta isoform expression found in colon carcinoma may signal an escape from the senescence barrier during the progression from adenoma to carcinoma.