The archetypal human tumor suppressor p53 is considered to have unique transactivation properties. The assumption is based on the fact that additionally identified human p53 isoforms lack transcriptional activity. However, we provide evidence for the existence of an alternatively spliced p53 isoform (Deltap53) that exerts its transcriptional activity independent from p53. In contrast to p53, Deltap53 transactivates the endogenous p21 and 14-3-3sigma but not the mdm2, bax, and PIG3 promoter. Cell cycle studies showed that Deltap53 displays its differential transcriptional activity only in damaged S phase cells. Upon activation of the ATR-intra-S phase checkpoint, Deltap53, but not p53, transactivates the Cdk inhibitor p21. Induction of p21 results in downregulation of cyclin A-Cdk activity and accordingly attenuation of S phase progression. Data demonstrate that the Deltap53-p21-cyclin A-Cdk pathway is crucial to facilitate uncoupling of repair and replication events, indicating that Deltap53 is an essential element of the ATR-intra-S phase checkpoint.
We demonstrate that wild-type p53 inhibits homologous recombination. To analyze DNA substrate specificities in this process, we designed recombination experiments such that coinfection of simian virus 40 mutant pairs generated heteroduplexes with distinctly unpaired regions. DNA exchanges producing single C-T and A-G mismatches were inhibited four-to sixfold more effectively than DNA exchanges producing G-T and A-C single-base mispairings or unpaired regions of three base pairs comprising G-T/A-C mismatches. p53 bound specifically to three-stranded DNA substrates, mimicking early recombination intermediates. The K D values for the interactions of p53 with three-stranded substrates displaying differently paired and unpaired regions reflected the mismatch base specificities observed in recombination assays in a qualitative and quantitative manner. On the basis of these results, we would like to advance the hypothesis that p53, like classical mismatch repair factors, checks the fidelity of homologous recombination processes by specific mismatch recognition.p53 germ line mutations are associated with a deficit to maintain genomic stability along with an increase of spontaneous gene amplification rates (17,52,93), thereby accelerating the multistep process of tumor progression (81). This phenotype has been explained by the loss of p53 cell cycle checkpoint control (38,39,46). DNA damage (38, 39, 60) and suboptimal growth situations, such as an increase of oxygen radicals (28) or ribonucleotide depletion (50), are signals for p53-mediated accumulation and functional activation (54,68). Depending on the cell type, p53 induces cell cycle arrest or apoptosis predominantly via transcriptional transactivation of genes coding for the cyclin-dependent kinase inhibitor p21/WAF1/CIP1/SDI1 (21, 23) or the apoptotic factor Bax (56). As a consequence, cells are unable to replicate their DNA under conditions which may lead or may have led to chromosome breaks (3), thereby preventing the manifestation and aggravation of genomic lesions in S phase. Strikingly, the same molecular signal triggering the DNA damage response by p53, namely, DNA strand breaks (60), also initiates V(D)J recombination (79), meiotic recombination (27), recombination repair (75), and gene amplification (19) events. There is evidence for an at least indirect involvement of p53 in V(D)J recombination, as ␥ irradiation can rescue rearrangement at multiple T-cell receptor loci by a p53-dependent bypass mechanism in scid mice (2, 12). A role for p53 in meiotic recombination has been postulated from the observation that p53 mRNA expression in testes of mice is high and specific for spermatocytes in zygotene to pachytene, the meiotic stages at which homologous chromosomes synapse for genetic exchange (65, 69). Intriguingly, the mitotic checkpoint factor Atm, the product of the gene mutated in patients with ataxia telangiectasia (66), is also found in spermatocytes of meiosis I. Atm belongs to the family of phosphatidylinositol 3-kinase-like protein kinases which, li...
In this study, we characterize the molecular and functional features of a novel protein called SPOC1. SPOC1 RNA expression was previously reported to be highest in highly proliferating tissues and increased in a subset of ovarian carcinoma patients, which statistically correlated with poor prognosis and residual disease. These observations implied that SPOC1 might play a role in cellular proliferation and oncogenesis. Here we show that the endogenous SPOC1 protein is labile, primarily chromatin associated and its expression as well as localization are regulated throughout the cell cycle. SPOC1 is dynamically regulated during mitosis with increased expression levels and biphasic localization to mitotic chromosomes indicating a functional role of SPOC1 in mitotic processes. Consistent with this postulate, SPOC1 siRNA knockdown experiments resulted in defects in mitotic chromosome condensation, alignment and aberrant sister chromatid segregation. Finally, we have been able to show, using micrococcal nuclease (MNase) chromatin-digestion assays that SPOC1 expression levels proportionally influence the degree of chromatin compaction. Collectively, our findings show that SPOC1 modulates chromatin structure and that tight regulation of its expression levels and subcellular localization during mitosis are crucial for proper chromosome condensation and cell division.
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