The ribosomal protein (RP)-MDM2 interaction is a p53 response pathway critical for preventing oncogenic c-MYC-induced tumorigenesis. To investigate whether the RP-MDM2-p53 pathway is a broad anti-oncogenic mechanism, we crossed mice bearing an MDM2C305F mutation, which disrupts RPL11 binding to MDM2, with mice expressing an oncogenic HrasG12V transgene. Interestingly, the MDM2C305F mutant mice, which are hypersensitive to c-MYC-induced tumorigenesis, are not hypersensitive to oncogenic HrasG12V-induced tumorigenesis. Unlike c-MYC, which induces expression of RPL11, RAS overexpression leads to an increase in RPL23 mRNA and protein while RPL11 expression remains unchanged. The induction of RPL23 involves both MEK and PI3K signaling pathways and requires mTOR function. Increased expression of RPL23, which maintains binding to MDM2C305F mutant, correlates with increased p53 expression in MDM2C305F cells. Furthermore, RAS overexpression can induce p53 in the absence of p19ARF, and the induction can be abolished by down-regulation of RPL23. Thus, while the RPL11-MDM2-p53 pathway coordinates with the p19ARF-MDM2-p53 pathway against oncogenic c-MYC-induced tumorigenesis, the RPL23-MDM2-p53 pathway coordinates with the p19ARF-MDM2-p53 pathway against oncogenic RAS-induced tumorigenesis.

Inactivation of the adenomatous polyposis coli (APC) tumor suppressor is frequently found in colorectal cancer. Loss of APC function results in deregulation of the Wnt/β-catenin signaling pathway causing overexpression of the c-MYC oncogene. In lymphoma, both p19ARF and ribosomal proteins RPL11 and RPL5 respond to c-MYC activation to induce p53. Their role in c-MYC-driven colorectal carcinogenesis is unclear, as p19ARF deletion does not accelerate APC loss-triggered intestinal tumorigenesis. To determine the contribution of the RP-MDM2-p53 pathway to APC loss-induced tumorigenesis, we crossed mice bearing MDM2C305F mutation, which disrupts RPL11- and RPL5-MDM2 binding, with Apcmin/+ mice, which are prone to intestinal tumor formation. Interestingly, loss of RP-MDM2 binding significantly accelerated colorectal tumor formation while having no discernable effect on small intestinal tumor formation. Mechanistically, APC loss leads to overexpression of c-MYC, RPL11 and RPL5 in mouse colonic tumor cells irrespective of MDM2C305F mutation. However, notable p53 stabilization and activation were observed only in Apcmin/+;Mdm2+/+ but not Apcmin/+;Mdm2C305F/C305F colon tumors. These data establish that the RP-MDM2-p53 pathway, in contrast to the p19ARF-MDM2-p53 pathway, is a critical mediator of colorectal tumorigenesis following APC loss.

It is evident that p53 activity is critical for tumour prevention and stress response through its transcriptional activation of genes affecting cellular senescence, apoptosis, cellular metabolism, and DNA repair. The regulation of p53 is highly complex, and MDM2 and MDMX are thought to be critical for deciding the fate of p53, both through inhibitory binding and post-translational modification. Many mouse models have been generated to study the regulation of p53 in vivo, and they have altered our interpretations of how p53 is regulated by MDM2 and MDMX. Although MDM2 is absolutely required for p53 regulation, certain functions are dispensable under unstressed conditions, including the ability of MDM2 to degrade p53. MDMX, on the other hand, may only be required in select situations, like embryogenesis. These models have also clarified how cellular stress signals modify the p53-inhibiting activities of MDM2 and MDMX in vivo. It is clear that more work will need to be performed to further understand the contexts for each of these signals and the requirements of various MDM2 and MDMX functions. Here, we will discuss what we have learned from mouse modelling of MDM2 and MDMX and underscore the ways in which these models could inform future therapies.

The MDM2 RING domain harbors E3 ubiquitin ligase activity critical for regulating the degradation of tumor suppressor p53, which controls many cellular pathways. The MDM2 RING domain also is required for an interaction with MDMX. Mice containing a substitution in the MDM2 RING domain, MDM2, disrupting MDM2 E3 function and the MDMX interaction, die during early embryogenesis that can be rescued by p53 deletion. To investigate whether MDM2, which retains p53 binding, has p53-suppressing activity, we generated ; mice, in which we replaced the endogenous alleles with an inducible allele, and compared survival with that of similarly generated ; mice. Adult -null mice died ∼7 days after tamoxifen-induced p53 activation, indicating that in the absence of MDM2, MDMX cannot suppress p53. Surprisingly, ; mice died ∼5 days after tamoxifen injection, suggesting that p53 activity is higher in the presence of MDM2 than in the absence of MDM2. Indeed, in MDM2-expressing mouse tissues and embryonic fibroblasts, p53 exhibited higher transcriptional activity than in those expressing no MDM2 or no MDM2 and MDMX. This observation indicated that MDM2 not only is unable to suppress p53 but may have gained the ability to enhance p53 activity. We also found that p53 acetylation, a measure of p53 transcriptional activity, was higher in the presence of MDM2 than in the absence of MDM2. These results reveal an unexpected role of MDM2 in enhancing p53 activity and suggest the possibility that compounds targeting MDM2 RING domain function could produce even more robust p53 activation.

<div>Abstract<p>The ribosomal protein (RP)–MDM2 interaction is a p53 response pathway critical for preventing oncogenic c-MYC–induced tumorigenesis. To investigate whether the RP-MDM2-p53 pathway is a broad antioncogenic mechanism, we crossed mice bearing an MDM2^C305F mutation, which disrupts RPL11 binding to MDM2, with mice expressing an oncogenic <i>Hras^G12V</i> transgene. Interestingly, the MDM2^C305F-mutant mice, which are hypersensitive to c-MYC–induced tumorigenesis, are not hypersensitive to oncogenic <i>Hras^G12V</i>-induced tumorigenesis. Unlike c-MYC, which induces expression of RPL11, RAS overexpression leads to an increase in RPL23 mRNA and protein whereas RPL11 expression remains unchanged. The induction of RPL23 involves both MEK and PI3K signaling pathways and requires mTOR function. Increased expression of RPL23, which maintains binding to MDM2^C305F mutant, correlates with increased p53 expression in MDM2^C305F cells. Furthermore, RAS overexpression can induce p53 in the absence of p19ARF, and the induction can be abolished by downregulation of RPL23. Thus, although the RPL11–MDM2–p53 pathway coordinates with the p19ARF–MDM2–p53 pathway against oncogenic c-MYC–induced tumorigenesis, the RPL23–MDM2–p53 pathway coordinates with the p19ARF–MDM2–p53 pathway against oncogenic RAS-induced tumorigenesis. <i>Cancer Res; 76(17); 5030–9. ©2016 AACR</i>.</p></div>