# Emerging Non-Canonical Functions and Regulation by p53: p53 and Stemess
## 4 Therapeutic Strategies to Restore _wt_p53 Function in CSCs
### RING Finger E3 Ligases and CSCs Maintenance  
Recent studies have discovered RING finger E3 ligases to be important contributors to cell stemness via regulation of self-renewal, differentiation and drug resistance displayed by CSCs. Inhibition of RING finger proteins may provide promising strategies for cancer treatment. RING finger proteins are not catalysts with active sites; instead, they facilitate ubiquitin transfer from an E2 directly to the substrate. Disrupting the RING structure or RING-E2 interface would represent viable routes for inhibitor development. Nutlin-3 is a small molecule antagonist, which binds to the  
Figure 4: Targeting of the p53 Network in cancer stem cells. (**A**) Restoring p53 transcriptional activity be preventing/restoring p53 exclusion from the nucleus; (**B**) targeting molecules to disrupt the p53-Mdm2 interaction to prevent p53 degradation; (**C**) restoring mutant p53 function by modifications; (**D**) targeting CSC pathways. amino terminus of Mdm2 and prevents binding to p53, leading to increased stability of p53 [236]. As a result of stabilizing p53, cancer cells are able to undergo classic p53-mediated cell cycle arrest and apoptosis. [PERSON] et al. (2014) [263] reviewed the regulation of CSCs by RING finger ubiquitin ligases and reported that SKP1-CUL1-F-box protein (SCF) E3s, CBL, BRACA1, MDM2 and von Hippel-Lindau tumor suppressor (VHL) are crucial in the regulation of cell-cycle progression, cell differentiation and CSC maintenance. As the guardian of the genome, p53 maintains cellular integrity by inhibiting the cell cycle and preventing the proliferation of damaged cells [264]. These processes are controlled via negative regulation of the tumor suppressor p53. There are many ubiquitin ligases that can regulate p53, Mdm2, Pirh2, Cop1, Trim24, ArfBP1, Carp1/2 and TOPORs. While some of these ligases are associated with CSCs, their role with respect to tissue-specific regulation of p53 or even stimuli-specific regulation of p53 remains to be determined. ESCs and early embryos endogenously express high levels of full-length p53 and remain viable upon removal of p53 [13; 265; 266; 267]. However, mice lacking Mdm2 succumb to early embryonic lethality and expire before implantation. Active p53 leads to embryonic lethality in Mdm2-deficient mice, but interestingly, the phenotype can be rescued by concomitant deletion of p53, demonstrating that negative regulation of p53 is essential during early embryogenesis and for ESC survival [268]. In HSCs and progenitors, loss of Mdm2 activity stabilizes p53 and impedes hematopoiesis via induction of cell cycle arrest, senescence and cell death [176]. Mdm2 is overexpressed in many high grade cancers with over a third present in tumors with _wt_p53 [269]. The MDM2 gene is transcriptionally regulated by p53, and alternative splicing can give rise to several Mdm2 transcript variants expressed in cancer. Mdm2 can also bind to tumor suppressor ARF with the interaction resulting in sequestration of Mdm2 in the nucleolus, where it cannot bind and degrade p53, therefore activating p53. Researchers have embarked on associations of Mdm2 with stemness in a variety of cellular processes. [PERSON] et al. [270] (2015) demonstrated that a combined inhibition of Mdm2 and Mdm2 stimulating the AKT/mTOR pathway using inhibitors ISA27 and FC85, respectively, enhanced glioblastoma multiforme (GBM) cell apoptosis and differentiation of CSCs. In MEFs, [PERSON] et al. (2016) [271] demonstrated that efficient iPSC generation is dependent on Mdm2 in absence of p53. While in hMSCs, Mdm2 depletion promoted cell differentiation and decreased clonogenic survival in cancer. Inactivation of the Polycomb Repressor Complex2 (PRC2) and EZH2 can induce many genes regulated by Mdm2. The complex formation of Mdm2 chromatin and EZH2 increased trimethylation and ubiquitination of histone 2 (lysine 27) and histone 2A (lysine 119; H2 AK119), respectively. Furthermore, the absence of Mdm2 and H2 AK119 E3 ligase Ring1B/RNF2 inhibited cell proliferation.