Abstract
BackgroundThe tumor suppressor p53 is a sequence-specific transcription factor that regulates an extensive network of coding genes, long non-coding RNAs and microRNAs, that establish intricate gene regulatory circuits influencing many cellular responses beyond the prototypical control of cell cycle, apoptosis and DNA repair.MethodsUsing bioinformatic approaches, we identified an additional group of candidate microRNAs (miRs) under direct p53 transcriptional control. To validate p53 family-mediated responsiveness of the newly predicted target miRs we first evaluated the potential for wild type p53, p63β and p73β to transactivate from p53 response elements (REs) mapped in the miR promoters, using an established yeast-based assay.ResultsThe REs found in miR-10b, -23b, -106a, -151a, -191, -198, -202, -221, -320, -1204, -1206 promoters were responsive to p53 and 8 of them were also responsive to p63β or p73β. The potential for germline p53 mutations to drive transactivation at selected miR-associated REs was also examined. Chromatin Immuno-Precipitation (ChIP) assays conducted in doxorubicin-treated MCF7 cells and HCT116 p53+/+ revealed moderate induction of p53 occupancy at the miR-202, -1204, -1206, -10b RE-containing sites, while weak occupancy was observed for the miR-23b-associated RE only in MCF7 cells. RT-qPCR analyses cells showed modest doxorubicin- and/or Nutlin-dependent induction of the levels of mature miR-10b, -23b, -151a in HCT116 p53+/+ and MCF7 cells. The long noncoding RNA PVT1 comprising miR-1204 and −1206 was weakly induced only in HCT116 p53+/+ cells, but the mature miRs were not detected. miR-202 expression was not influenced by p53-activating stimuli in our cell systems.ConclusionsOur study reveals additional miRs, particularly miR-10b and miR-151a, that could be directly regulated by the p53-family of transcription factors and contribute to the tuning of p53-induced responses.
Highlights
The tumor suppressor p53 is a sequence-specific transcription factor that regulates an extensive network of coding genes, long non-coding RNAs and microRNAs, that establish intricate gene regulatory circuits influencing many cellular responses beyond the prototypical control of cell cycle, apoptosis and DNA repair
The master regulator p53 is a prominent tumor suppressor gene, functioning in the cell as a tetrameric sequence-specific transcription factor, able to bind to two copies of a decameric sequence with the RRRCWWGYYY consensus representing the so called p53-response element (p53-response elements (REs)) [1]. p53 is known to be inducible in response to a large number of cellular stress signals that, besides genotoxic stress, include carbon
To confirm p53 responsiveness of the identified p53 REs we first applied a well-established quantitative reporter assay in the budding yeast Saccharomyces cerevisiae [37,43]
Summary
The tumor suppressor p53 is a sequence-specific transcription factor that regulates an extensive network of coding genes, long non-coding RNAs and microRNAs, that establish intricate gene regulatory circuits influencing many cellular responses beyond the prototypical control of cell cycle, apoptosis and DNA repair. The authors demonstrated that genes encoding miR-34 family cluster were direct transcriptional targets of p53 and that their induced expression levels upon genotoxic or oncogenic stress was dependent on p53 expression, both in vitro and in vivo. He et al identified the DNA sequences responsible for the p53 responsiveness of those miRs. A year later another group of miRs, (miR-192, -194 and −215) was identified as targets of p53 and their ability to increase the level of CDKN1A (p21CIP1) and to function as drivers of cell cycle arrest was established [17]. Jin et al surprisingly found that p53 directly induced the transcription of miR149*, which in turn can target the glycogen synthase kinase-3α mRNA, resulting in elevated expression of Mcl-1 and resistance to apoptosis in melanoma cells, providing a rational explanation for the poor ability of p53 to suppress melanoma progression [21]
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