The p53 Tumor Suppressor Is Stabilized by Inhibitor of Growth 1 (ING1) by Blocking Polyubiquitination

Abstract

The INhibitor of Growth tumor suppressors (ING1-ING5) affect aging, apoptosis, DNA repair and tumorigenesis. Plant homeodomains (PHD) of ING proteins bind histones in a methylation-sensitive manner to regulate chromatin structure. ING1 and ING2 contain a polybasic region (PBR) adjacent to their PHDs that binds stress-inducible phosphatidylinositol monophosphate (PtIn-MP) signaling lipids to activate these INGs. ING1 induces apoptosis independently of p53 but other studies suggest proapoptotic interdependence of ING1 and p53 leaving their functional relationship unclear. Here we identify a novel ubiquitin-binding domain (UBD) that overlaps with the PBR of ING1 and shows similarity to previously described UBDs involved in DNA damage responses. The ING1 UBD binds ubiquitin with high affinity (Kd∼100 nM) and ubiquitin competes with PtIn-MPs for ING1 binding. ING1 expression stabilized wild-type, but not mutant p53 in an MDM2-independent manner and knockdown of endogenous ING1 depressed p53 levels in a transcription-independent manner. ING1 stabilized unmodified and six multimonoubiquitinated forms of wild-type p53 that were also seen upon DNA damage, but not p53 mutants lacking the six known sites of ubiquitination. We also find that ING1 physically interacts with herpesvirus-associated ubiquitin-specific protease (HAUSP), a p53 and MDM2 deubiquitinase (DUB), and knockdown of HAUSP blocks the ability of ING1 to stabilize p53. These data link lipid stress signaling to ubiquitin-mediated proteasomal degradation through the PBR/UBD of ING1 and further indicate that ING1 stabilizes p53 by inhibiting polyubiquitination of multimonoubiquitinated forms via interaction with and colocalization of the HAUSP-deubiquitinase with p53.

Function of the ING Proteins in Cancer and Senescence

“…This prevents the deacetylation of p53 by hSIR2amd thus maintains p53 in an active state. As noted previously, ING1 also regulates p21 levels by inhibiting ubiquitinmediated proteosomal degradation of p53 through physically interacting with herpesvirusassociated ubiquitin-specifi c protease (HAUSP), a p53 and MDM2 deubiquitinase (Thalappilly et al 2011 ).…”

“…However, several other domains are found in subsets of the INGs. ING1b contains a partial bromodomain (PBD), whose function remains unknown, and a PCNA-interacting-protein (PIP) motif, which helps to mediate ING1b binding with the prolif-erating cell nuclear antigen (PCNA) in a highly DNA damage-inducible manner (Soliman and Riabowol 2007 ;Thalappilly et al 2011 ). In ING2 to ING5, a leucine zipper-like (LZL) motif occupies the regions comparable to those of the PIP and PBD of ING1.…”

“…The LZL was reported to affect ING functions in DNA repair and apoptosis. Lastly, ING1 and ING2 contain a UBD or ubiquitin-binding domain that was found to be crucial for stabilizing p53 by affecting its ubiquitin-mediated proteosomal degradation (Thalappilly et al 2011 ). This domain overlaps with the polybasic region (PBR) found at the C-termini of ING1 and ING2 that was determined to be necessary and suffi cient for the binding of stress-inducible phosphatidylinositol 5′-monophosphatesignalling lipids, resulting in the promotion of apoptosis ).…”

“…This post-translational modifi cation acts to increase the stability of ING1, thus, regulating the expression of cyclin B1 and proliferation in melanoma cells. ING1 has also been reported to be ubiquitinated as a mechanism of turnover via the proteasome, but it also binds ubiquitin through its UBD (Thalappilly et al 2011 ).…”

Hypoxic regulation of the expression of cell proliferation related genes in U87 glioma cells upon inhibition of IRE1 signaling enzyme

“…Reduced expression and rearrangement of this gene have been detected in various cancers [8,9]. Moreover, ING1 stabilizes p53 by inhibiting polyubiquitination [10]. KRT18 (keratin 18) contributes to decreased malignancy of non-small cell lung carcinoma and is directly regulated by EGR1 [11].…”