Fine-tuning the p53 response to DNA damage: A new piece in the puzzle

Abstract

The tumor suppressor p531 is a master regulator of cell cycle checkpoint responses to DNA damage whose activity is tightly controlled. It functions as a transcription factor for genes regulating cell cycle progression and apoptotic cell death. Following DNA damage, p53 is stabilized and its transcriptional activity is markedly stimulated. Among the many factors that can modulate p53 activity, the RNA-binding protein hnRNP K (heterogeneous ribonucleoprotein K) was characterized as a key player in the p53-dependent response to DNA damage in human cells, acting as a transcriptional coactivator for p53.2 hnRNP K and p53 are indeed co-recruited to the promoters of p53-responsive genes and cooperate to elicit their activation. Interestingly, similar to p53, hnRNP K levels are regulated by the E3 ubiquitin ligase HDM2, which targets hnRNP K for ubiquitylation and proteasomal degradation.2 DNA damage triggers the dissociation of the hnRNP K-HDM2 complex, leading to hnRNP K stabilization. Because these events are dependent on the DNA damage checkpoint kinases ATM (ataxia telangiectasia mutated) and ATR (ATM- and Rad3-related),2 it was tempting to speculate that hnRNP K function in the DDR (DNA damage response) could be regulated by phosphorylation. This issue has been explored in a recent study published in Cell Cycle, where Moumen et al. showed that a phosphorylated form of hnRNP K can be detected with a phospho-ATM/ATR substrate antibody upon IR (ionizing radiation)-induced damage in human cells.3 Given that hnRNP K levels are elevated in an ATM-dependent manner in response to IR,2 the authors tested the possibility that this key upstream DNA damage checkpoint kinase could target hnRNP K. Using an ATM-specific inhibitor and siRNA-mediated depletion of the ATM kinase, they demonstrate that hnRNP K is phosphorylated in an ATM-dependent manner after IR. They also identify four S/T-Q ATM consensus target motifs in the hnRNP K sequence (S121, T174, T390, T440) that can serve as ATM phosphorylation sites. Similar phosphorylation events may be mediated by ATR following UV irradiation, because hnRNP K is stabilized in an ATR-dependent manner in this context.2 Next, Moumen et al. examined the functional relevance of such phosphorylation of hnRNP K in the DDR. Taking advantage of a phospho-deficient mutant where the four S/T sites are mutated to alanines, they established that phosphorylation of hnRNP K is critical for its dissociation from HDM2 and subsequent stabilization in response to IR. It is not yet clear whether the ATM-dependent phosphorylation of hnRNP K directly inhibits its interaction with HDM2, because how hnRNP K interacts with HDM2 is currently unknown. Importantly, cells expressing the phospho-deficient hnRNP K protein also display impaired recruitment of p53 to its target gene p21Waf1 and defective stimulation of p53 transcriptional activity. Together, these findings support a model where ATM-mediated phosphorylation protects hnRNP K from proteasomal degradation in response to damage, allowing it function as a coactivator for p53 (Fig. 1). Identifying phosphatases able to reverse hnRNP K phosphorylation would help define how this response is switched off after damage. Figure 1. Control of hnRNP K by phosphorylation: Integrating DNA damage signals to fine-tune p53-dependent transcriptional responses. Remarkably, such phospho-dependent control of hnRNP K levels in response to DNA damage mirrors the upregulation of p53, whose phosphorylation by ATM also leads to its stabilization upon dissociation from HDM2. This striking parallel between two proteins collaborating in the DDR reveals the tight control that ATM exerts on p53 transcriptional activity. By achieving the same goal via two converging pathways, ATM imposes a double lock on cell cycle checkpoint responses (Fig. 1). These observations should be considered in light of other DNA damage-dependent modifications on hnRNP K that control p53-dependent transcription, including sumoylation and methylation.4-6 It will be of major interest to identify potential cross-talk and/or interference between these modifications to shed light on how they collectively contribute to the fine-tuning of hnRNP K activity in response to damage. It would also be important to evaluate whether ATM-dependent phosphorylation effects other functions of hnRNP K in RNA metabolism, including its ability to repress p53-target genes via an interaction with the p53-induced large intergenic noncoding RNA lincRNA-p21.7 Finally, the critical role of ATM phospho-target sites in regulating hnRNP K protein levels opens up the possibility that mutations of these residues could be instrumental in driving the upregulation of hnRNP K frequently observed in human tumors.