LAUSR

p53 tumor suppressor, the most mutated gene in human cancer, is negatively regulated by MDM2 and MDM4 proteins. Two distinct mechanisms of MDM2/MDM4 regulation of p53 include, one, suppression of p53 transactivation activity by direct interaction of MDM2/MDM4 with p53 protein, thus blocking its transactivation interface and two, targeting p53 to proteasomal degradation by MDM2 E3 ligase activity. However, the importance of these mechanisms in normal p53 regulation in vivo has remained controversial. To genetically separate these two mechanisms of p53 regulation we generated a novel Mdm2 mutant mouse (Mdm2L466A) that lacks E3 ligase activity but retains the ability of Mdm2 to heterodimerize with Mdm4 and inhibit p53 transactivation. Homozygous Mdm2L466A mice are embryonic lethal due to dysregulated p53 activity, thus demonstrating Mdm2 E3 ligase mediated regulation of p53 is essential during embryonic development. Additionally, we uncovered novel p53-independent functions of MDM2 E3 ligase in cell cycle regulation and genome integrity in cells. Cells lacking MDM2 E3 ligase activity have defective G2-M transition during cell cycle and developed elevated levels of aneuploidy regardless of p53 status. This study unequivocally demonstrates the requirement of Mdm2 E3 ligase activity for normal regulation of p53 and uncovers previously unknown p53-independent role of Mdm2 in cell cycle regulation and maintaining genome integrity. A current therapeutic approach involves blocking MDM2-p53 interaction to promote p53 tumor suppression function in cancer cells. Our findings suggest that this approach alone is insufficient because MDM2 has other potential oncogenic mechanisms independent of its role as negative regulator of p53. Hence, targeting MDM2 E3 ligase activity is likely to be more effective cancer therapy as it addresses both p53-dependent and p53-independent oncogenic mechanisms of MDM2. Citation Format: Meenalakshmi Chinnam, Rati Lama, Chao Xu, Xiaojing Zhang, Carlos Cedeno, Yanqing Wang, Aimee B. Stablewski, David W. Goodrich, Xinjiang Wang. Requirement of MDM2 E3 ligase activity for regulating p53 during normal development, cell cycle regulation and genome integrity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2606.