NIR is a novel INHAT repressor that modulates the transcriptional activity of p53.

Most transcriptional repression pathways depend on the targeted deacetylation of histone tails. In this report, we characterize NIR, a novel transcriptional corepressor with inhibitor of histone acetyltransferase (INHAT) activity. NIR (Novel INHAT Repressor) is ubiquitously expressed throughout embryonic development and adulthood. NIR is a potent transcriptional corepressor that is not blocked by histone deacetylase inhibitors and is capable of silencing both basal and activator-driven transcription. NIR directly binds to nucleosomes and core histones and prevents acetylation by histone acetyltransferases, thus acting as a bona fide INHAT. Using a tandem affinity purification approach, we identified the tumor suppressor p53 as a NIR-interacting partner. Association of p53 and NIR was verified in vitro and in vivo. Upon recruitment by p53, NIR represses transcription of both p53-dependent reporters and endogenous target genes. Knock-down of NIR by RNA interference significantly enhances histone acetylation at p53-regulated promoters. Moreover, p53-dependent apoptosis is robustly increased upon depletion of NIR. In summary, our findings describe NIR as a novel INHAT that plays an important role in the control of p53 function.

No anti-apoptotic effects of single copies of mutant p53 genes in drug-treated tumor cells.

Some mutant forms of the p53 tumor suppressor have been documented to exert novel oncogenic functions including the increase of tumorigenicity, metastatic potential, genomic instability and therapy resistance of tumor cells. The latter has been suggested to be caused, primarily, by inhibition of apoptosis and, in part, through the activation of genes by mutant p53 whose products can counteract drug activities. Recently described in this context was the dUTPase, which may confer resistance to fluoropyrimidine drugs such as 5-fluorouracil (5-FU). We report here findings that call in question the existence of a direct anti-apoptotic effect of mutant p53. Wild-type p53-negative human fibroblasts, and Saos-2, H1299 and HCT116 tumor cells, treated with adriamycin, etoposide, cisplatin or 5-FU, failed to show apoptosis resistance when retrovirally bulk-infected to express the p53 mutants 175H or 273H at levels observed in naturally mutant p53-producing tumor cells. Furthermore, dUTPase gene expression was not stimulated by mutant p53, but instead by cellular events that involve DNA synthesis. We interpret the combined available data to suggest that much of the anti-apoptotic effect of mutant p53 is indirect and secondary to DNA-damaging and/or repair-interfering effects of these proteins.

Mitochondrial p53 levels parallel total p53 levels independent of stress response in human colorectal carcinoma and glioblastoma cells.

p53 can eliminate damaged cells through the induction of mitochondria-mediated apoptosis. Recent observations have provided strong evidence that a fraction of total p53 translocates to mitochondria specifically in response to a death stimulus. Unexpectedly, mutant p53, which is expressed at much higher levels than wild type in unstressed cells, is apparently always present at the mitochondria, independent of apoptotic signal. This prompted us to ask whether cell lines with intact p53-dependent apoptosis and cell cycle arrest pathways exist in which the mitochondrial localization of wild-type p53, like that of mutant, is independent of a death stimulus and instead, correlates with the total p53 levels. Here, we document that human HCT116 colorectal carcinoma cells treated with adriamycin or 5-fluorouracil (5FU) can accumulate total p53 to equally high levels, and mitochondrial p53 to proportionate levels, although only 5FU treatment provoked p53-dependent apoptosis. Along the same line, HCT116 derivatives with increased basal p53 levels, and glioblastoma cells with a doxycycline-inducible p53, also revealed proportionate mitochondrial p53 levels, and even unstressed HCT116 cells had some p53 located at the mitochondria. Finally, mitochondrial and total p53 showed distinct post-translational modifications. Thus, cell lines exist in which the mitochondrial p53 levels parallel total levels independent of apoptosis.