The novel p53 target gene IRF2BP2 participates in cell survival during the p53 stress response

The tumor suppressor p53 contributes to the cellular fate after genotoxic insults, mainly through the regulation of target genes, thereby allowing e.g. repair mechanisms resulting in cell survival or inducing apoptosis. Unresolved so far is the issue, which exact mechanisms lead to one or the other cellular outcome. Here, we describe the interferon regulatory factor-2-binding protein-2 (IRF2BP2) as a new direct target gene of p53, influencing the p53-mediated cellular decision. We show that upregulation of IRF2BP2 after treatment with actinomycin D (Act.D) is dependent on functional p53 in different cell lines. This occurs in parallel with the down-regulation of the interacting partner of IRF2BP2, the interferon regulatory factor-2 (IRF2), which is known to positively influence cell growth. Analyzing the molecular functions of IRF2BP2, it appears to be able to impede on the p53-mediated transactivation of the p21- and the Bax-gene. We show here that overexpressed IRF2BP2 has an impact on the cellular stress response after Act.D treatment and that it diminishes the induction of apoptosis after doxorubicin treatment. Furthermore, the knockdown of IRF2BP2 leads to an upregulation of p21 and faster induction of apoptosis after doxorubicin as well as Act.D treatment.     

Measuring a cell's response to stress: the p53 pathway

The characterization of complex cellular responses to diverse stimuli can be studied by the use of emerging chip-based technologies.     

MDMX regulation of p53 response to ribosomal stress

Ribosomal stress such as disruption of rRNA biogenesis activates p53 by release of ribosomal proteins from the nucleoli, which bind to MDM2 and inhibit p53 degradation. We found that p53 activation by ribosomal stress requires degradation of MDMX in an MDM2-dependent fashion. Tumor cells overexpressing MDMX are less sensitive to actinomycin D-induced growth arrest due to formation of inactive p53-MDMX complexes. Knockdown of MDMX increases sensitivity to actinomycin D, whereas MDMX overexpression abrogates p53 activation and prevents growth arrest. Furthermore, MDMX expression promotes resistance to the chemotherapeutic agent 5-fluorouracil (5-FU), which at low concentrations activates p53 by inducing ribosomal stress without significant DNA damage signaling. Knockdown of MDMX abrogates HCT116 tumor xenograft formation in nude mice. MDMX overexpression does not accelerate tumor growth but increases resistance to 5-FU treatment in vivo. Therefore, MDMX is an important regulator of p53 response to ribosomal stress and RNA-targeting chemotherapy agents.     

Inhibition of nucleolar stress response by Sirt1: A potential mechanism of acetylation‐independent regulation of p53 accumulation

The mammalian Sirt1 deacetylase is generally thought to be a nuclear protein, but some pilot studies have suggested that Sirt1 may also be involved in orchestrating nucleolar functions. Here, we show that nucleolar stress response is a ubiquitous cellular reaction that can be induced by different types of stress conditions, and Sirt1 is an endogenous suppressor of nucleolar stress response. Using stable isotope labeling by amino acids in cell culture approach, we have identified a physical interaction of between Sirt1 and the nucleolar protein nucleophosmin, and this protein–protein interaction appears to be necessary for Sirt1 inhibition on nucleolar stress, whereas the deacetylase activity of Sirt1 is not strictly required. Based on the reported prerequisite role of nucleolar stress response in stress‐induced p53 protein accumulation, we have also provided evidence suggesting that Sirt1‐mediated inhibition on nucleolar stress response may represent a novel mechanism by which Sirt1 can modulate intracellular p53 accumulation independent of lysine deacetylation. This process may represent an alternative mechanism by which Sirt1 regulates functions of the p53 pathway.     

Hzf Determines cell survival upon genotoxic stress by modulating p53 transactivation

A critical unresolved issue about the genotoxic stress response is how the resulting activation of the p53 tumor suppressor can lead either to cell-cycle arrest and DNA repair or to apoptosis. We show here that hematopoietic zinc finger (Hzf), a zinc-finger-containing p53 target gene, modulates p53 transactivation functions in an autoregulatory feedback loop. Hzf is induced by p53 and binds to its DNA-binding domain, resulting in preferential transactivation of proarrest p53 target genes over its proapoptotic target genes. Thus, p53 activation results in cell-cycle arrest in Hzf wild-type MEFs, while in Hzf(-/-) MEFs, apoptosis is induced. Exposure of Hzf null mice to ionizing radiation resulted in enhanced apoptosis in several organs, as compared to in wild-type mice. These findings provide novel insights into the regulation of p53 transactivation function and suggest that Hzf functions as a key player in regulating cell fate decisions in response to genotoxic stress.     

The p53-inducible E3 ubiquitin ligase p53RFP induces p53-dependent apoptosis

Recently, it has been shown that really interesting new gene (RING)-in between ring finger (IBR)-RING domain-containing proteins, such as Parkin and Parc, are E3 ubiquitin ligases and are involved in regulation of apoptosis. In this report, we show that p53-inducible RING-finger protein (p53RFP), a p53-inducible E3 ubiquitin ligase, induces p53-dependent but caspase-independent apoptosis. p53RFP contains an N-terminal RING-IBR-RING domain and an uncharacterized, evolutionally highly conserved C-terminal domain. p53RFP interacts with E2 ubiquitin-conjugating enzymes UbcH7 and UbcH8 but not with UbcH5, and this interaction is mediated through the RING-IBR-RING domain of p53RFP. Interestingly, the conserved C-terminal domain of p53RFP is required and sufficient for p53RFP-mediated apoptosis, suggesting p53RFP-mediated apoptosis does not require its E3 ubiquitin ligase activity. Together with a recent report showing that p53RFP is involved in ubiquitination and degradation of p21, a p53 downstream protein promoting growth arrest and antagonizing apoptosis, our findings suggest that p53RFP is involved in switching a cell from p53-mediated growth arrest to apoptosis.