Gomes NP, Espinosa JMGene-specific repression of the p53 target gene PUMA via intragenic CTCF-Cohesin binding. Genes Dev 24:1022-1034

Howard Hughes Medical Institute, Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, Colorado 80309, USA.
Genes & development (Impact Factor: 10.8). 05/2010; 24(10):1022-34. DOI: 10.1101/gad.1881010
Source: PubMed


The p53 transcriptional program orchestrates alternative responses to stress, including cell cycle arrest and apoptosis, but the mechanism of cell fate choice upon p53 activation is not fully understood. Here we report that PUMA (p53 up-regulated modulator of apoptosis), a key mediator of p53-dependent cell death, is regulated by a noncanonical, gene-specific mechanism. Using chromatin immunoprecipitation assays, we found that the first half of the PUMA locus (approximately 6 kb) is constitutively occupied by RNA polymerase II and general transcription factors regardless of p53 activity. Using various RNA analyses, we found that this region is constitutively transcribed to generate a long unprocessed RNA with no known coding capacity. This permissive intragenic domain is constrained by sharp chromatin boundaries, as illustrated by histone marks of active transcription (histone H3 Lys9 trimethylation [H3K4me3] and H3K9 acetylation [H3K9Ac]) that precipitously transition into repressive marks (H3K9me3). Interestingly, the insulator protein CTCF (CCCTC-binding factor) and the Cohesin complex occupy these intragenic chromatin boundaries. CTCF knockdown leads to increased basal expression of PUMA concomitant with a reduction in chromatin boundary signatures. Importantly, derepression of PUMA upon CTCF depletion occurs without p53 activation or activation of other p53 target genes. Therefore, CTCF plays a pivotal role in dampening the p53 apoptotic response by acting as a gene-specific repressor.

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Available from: Joaquín M Espinosa, Mar 20, 2015
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    • "Upon p53 activation, resulting in functional PUMA mRNA accumulation confirmed the results of our RT-qPCR analyses (Figure 10(a)). These observations in correlation with the results of other in vitro studies [80] suggested that PUMA a major effector of p53-mediated cell death may play an important role in in vitro regulator of apoptosis when it overexpressed. Moreover the results of the overexpression of p53 mRNA and PUMA mRNA (Figure 10(a)) taken together corroborated the findings expressed by Tsai and Barton [82], suggesting apparently that the branch of apoptosis put in evidence here was the intrinsic mitochondrial pathway. "
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    • "Of these, the PUMA RE has moderate transactivation potential based on its sequence features. Interestingly, the discovery of an insulator-type RNA-polymerase II stop site in proximity to exon 3 led to the proposal (82) that there is a post-transcriptional initiation mechanism for the regulation of PUMA expression, suggesting that in this case fine tuning of p53 affinity for the promoter may not be critical. "
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    • "Cohesin–DNA interactions are known to be influenced by loading factors (Scc2-Scc4) (34,35), transcription factors (e.g. mediator complex or CTCF) (11,36) and the DNA double-strand break repair machinery (9). Here we have shown that the core heterodimer alone (SMC1/3) is able to directly interact with DNA to drive a stepwise compaction process, and to mediate interactions between separate DNA molecules. "
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