MDM4/HIPK2/p53 cytoplasmic assembly uncovers coordinated repression of molecules with anti-apoptotic activity during early DNA damage response

Abstract

The p53 inhibitor, MDM4 (MDMX) is a cytoplasmic protein with p53-activating function under DNA damage conditions. Particularly, MDM4 promotes phosphorylation of p53 at Ser46, a modification that precedes different p53 activities. We investigated the mechanism by which MDM4 promotes this p53 modification and its consequences in untransformed mammary epithelial cells and tissues. In response to severe DNA damage, MDM4 stimulates p53Ser46P by binding and stabilizing serine–threonine kinase HIPK2. Under these conditions, the p53-inhibitory complex, MDM4/MDM2, dissociates and this allows MDM4 to promote p53/HIPK2 functional interaction. Comparative proteomic analysis of DNA damage-treated cells versus -untreated cells evidenced a diffuse downregulation of proteins with anti-apoptotic activity, some of which were targets of p53Ser46P/HIPK2 repressive activity. Importantly, MDM4 depletion abolishes the downregulation of these proteins indicating the requirement of MDM4 to promote p53-mediated transcriptional repression. Consistently, MDM4-mediated HIPK2/p53 activation precedes HIPK2/p53 nuclear translocation and activity. Noteworthy, repression of these proteins was evident also in mammary glands of mice subjected to γ-irradiation and was significantly enhanced in transgenic mice overexpressing MDM4. This study evidences the flexibility of MDM2/MDM4 heterodimer, which allows the development of a positive activity of cytoplasmic MDM4 towards p53-mediated transcriptional function. Noteworthy, this activity uncovers coordinated repression of molecules with shared anti-apoptotic function which precedes active cell apoptosis and that are frequently overexpressed and/or markers of tumour phenotype in human cancer.

MDM4 actively restrains cytoplasmic mTORC1 by sensing nutrient availability

“…This MDM4 function is in agreement with its pro-apoptotic activity under DNA damage and support a model of MDM4 with anti-oncogenic activities in stress conditions [43]. Furthermore, these data together with the previous report of MDM4 functioning as a bridge for phosphorylation of p53 [7], contribute to define MDM4 as a cytoplasmic scaffold ready to sense different stimuli – i.e. DNA damage, cell starvation – and to accordingly regulate cell growth by recruiting different partners.…”

Targeting the MDM2/MDM4 Interaction Interface as a Promising Approach for p53 Reactivation Therapy

“…The observation that the preferred response elicited by Pep3 is apoptosis is also in agreement with recent data from our and other labs that reported impairment of MDM4/MDM2 heterodimer function during apoptosis (22,(39)(40)(41). Importantly, this observation highlights the potential therapeutic value of targeting the MDM2/MDM4 heterodimers.…”

The Double Role of p53 in Cancer and Autoimmunity and Its Potential as Therapeutic Target

“…In agreement with these data, the lethality of the mdm4 and mdm2 KO mice is accomplished by an apoptotic outcome in the mdm2 KO and mostly by a growth arrest response in the mdm4 KO [23,24,25]. The dissociation of MDM4 from MDM2 increases the activation of p53 whereas the association of MDM2 to MDM4 counteracts the proapoptotic activity of the last [34]. …”