Activating p53 function by targeting RLIP

Abstract

Aberrations in RLIP, p53, and PKCα represent essentially the entire spectrum of all human neoplasms. Elevated PKCα expression, failure of the cell cycle checkpoint (p53 dysfunction), and abnormal glutathione (GSH) metabolism are fundamental hallmarks of carcinogenesis and drug/radiation resistance. However, a lack of investigations into the interactions between these important regulatory nodes has fundamentally limited our understanding of carcinogenesis and the development of effective interventions for cancer prevention and therapy. Loss of p53, perhaps the most powerful tumor suppressor gene, predisposes rodents to spontaneous cancer and humans to familial, as well as acquired, cancers. Until recently, no genetic manipulation of any oncogene had been reported to abrogate spontaneous carcinogenesis in p53^-/- rodent models. However, the overexpression of RLIP, a GSH-electrophile conjugate (GS-E) transporter, has been found to enhance cancer cell proliferation and confer drug/radiation resistance, whereas its depletion causes tumor regression, suggesting its importance in cancer and drug/radiation resistance. Indeed, RLIP is an essential effector of p53 that is necessary for broad cancer-promoting epigenetic remodeling. Interestingly, through a haploinsufficiency mechanism, the partial depletion of RLIP in p53^-/- mice provides complete protection from neoplasia. Furthermore, RLIP^-/- mice exhibit altered p53 and PKCα function, marked deficiency in clathrin-dependent endocytosis (CDE), and almost total resistance to chemical carcinogenesis. Based on these findings, in this review, we present a novel and radical hypothesis that expands our understanding of the highly significant cross-talk between p53, PKCα, and GSH signaling by RLIP in multiple tumor models.

Keywords: Drug resistance; Glutathione-electrophile conjugate; Mercapturic acid pathway; Metastasis; PKCα; RLIP; RalBP1; Therapeutics; p53.

Figure 1

(A) Oxidative metabolism of PUFAs. A model for the mechanism by which RLIP controls signaling by regulating cellular levels of HNE and its metabolites (GS-HNE and GS-DHN). Endogenous electrophilic compounds, especially those derived from essential PUFAs, are metabolized through a GSH-linked process; the rate of this metabolism controls receptor–ligand signaling. (B) Protein–protein complexes in which RLIP has been identified. A proposed gene signature and network regulated by p53 and RLIP. RNA-seq gene expression data from p53^−/− and RLIP^+/− mice were normalized to wild-type expression, and genes that were differentially expressed were filtered at −log (p-value) <7. This analysis yielded a set of 23 genes that were significantly altered in opposing directions in p53^−/− vs. RLIP^−/− mice. An overlay of data from RLIP^−/− mice (C) and RLIP antisense (R508)-treated p53^−/− mice (E) showed a pattern in which the deviations from wild-type expression typically observed in p53^−/− mice (D) were normalized upon RLIP depletion. We predict that all mice with low RLIP expression will be cancer-resistant and exhibit the pattern seen in (C). Mice with normal or high RLIP levels will show a pattern that is determined by p53 or HSF. Balanced losses in RLIP and p53 should yield wild-type cancer susceptibility and a wild-type gene signature.

Figure 2. Genetic alterations of RLIP, HSF1, and p53 in cancer.

Genomic data from TCGA was queried using cBioportal for copy number alterations across 26 human cancers. The results (A) show total cases and genetic alterations in either gene. Case-wise alterations (mutation: green; truncation: black; deletion: blue; amplification: red; no change: gray) are represented for the 6,171 cases analyzed (B). Significant and notable findings from additional queries to determine the co-occurrence or exclusivity of these alterations are also presented (C), and a summary of our interpretation is given below.

Figure 3. A hypothetical model for the role of the RLIP–p53–Hsf1 interaction on carcinogenesis and DNA methylation.

H=HSF1; P=p53; R=RLIP; −/− = homozygous knockout; +/− = heterozygous knockout; RH, PH, and PR are heterodimers; SpCa = spontaneous cancer; InCa = inducible cancer; Met = DNA methylation (Upper Panel). A signaling model is presented with putative inhibition (red); activation (green); mutual regulation (blue); and enzymatic signaling (purple) (Lower Panel).

Figure 4

Proposed mechanism of cancer prevention in p53^−/− mice by RLIP depletion.