Abstract
The tumour suppressor p53 is an important mediator of cell cycle arrest and apoptosis in response to DNA damage, acting mainly by transcriptional regulation of specific target genes. The exact details how p53 modulates this decision on a molecular basis is still incompletely understood. One mechanism of regulation is acetylation of p53 on lysine K120 by the histone-acetyltransferase Tip60, resulting in preferential transcription of proapoptotic target genes. PDCD5, a protein with reported pro-apoptotic function, has recently been identified as regulator of Tip60-dependent p53-acetylation. In an effort to clarify the role of PDCD5 upon DNA damage, we generated cell lines in which PDCD5 expression was conditionally ablated by shRNAs and investigated their response to genotoxic stress. Surprisingly, we failed to note a rate-limiting role of PDCD5 in the DNA damage response. PDCD5 was dispensable for DNA damage induced apoptosis and cell cycle arrest and we observed no significant changes in p53 target gene transcription. While we were able to confirm interaction of PDCD5 with p53, we failed to do so for Tip60. Altogether, our results suggest a role of PDCD5 in the regulation of p53 function but unrelated to cell cycle arrest or apoptosis, at least in the cell types investigated.
Highlights
The tumour suppressor p53 is an important mediator of cell cycle arrest and apoptosis in response to DNA damage, acting mainly by transcriptional regulation of specific target genes
These findings are in line with a modulatory role on p53 activity, structural analysis and studies using the recombinant proteins suggested that the N-terminal domain of programmed cell death 5 (PDCD5) can exert a pro-death role on its own that may be independent from its proposed modulatory activity of p5330
In an effort to study the role of PDCD5 in the DNA damage response, we first investigated the transcriptional regulation of PDCD5 in Hct[116] colon carcinoma, A549 lung carcinoma and U2OS osteosarcoma cells
Summary
The tumour suppressor p53 is an important mediator of cell cycle arrest and apoptosis in response to DNA damage, acting mainly by transcriptional regulation of specific target genes. After DNA damage activates the DDR machinery, p53 is phosphorylated at several amino acids[11,12] These posttranslational modifications prevent its recognition and subsequent degradation by the E3 ubiquitin ligase MDM2 and enable p53 to translocate to the nucleus and induce transcription of its target genes. Acetylation of p53 on lysine (K) 120 has been shown to enable transcription of proapoptotic target genes like Puma or Bax[13,14] This modification is carried out by the histone acetyltransferase (HAT) Tip[60], which is activated in response to DNA damage by methylated histone H315. PDCD5 is transcriptionally upregulated in response to genotoxic stress and translocates to the nucleus where it is part of a complex containing Tip[60] and p5316,23, thereby enhancing the HAT activity of Tip[60] This in turn is thought to lead to increased K120 acetylation and increased transcription of proapoptotic genes promoting cell death. To explore the role of PDCD5 in cell death signalling in more detail we generated a PDCD5-specific antiserum that recognizes endogenous mouse and human PDCD5 protein as well as a series of human cancer cell lines where PDCD5 expression can be conditionally ablated by RNAi and studied their response to DNA damage
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