Identification of the RNAs that bind with MDM2 by high-throughput RIP-seq in MDA-MB-231 and MCF-7 cells

The E3 ligase MDM2 promotes tumor growth and progression by inducing ubiquitin-mediated degradation of P53 and other tumor suppressing proteins. Here, we identified an MDM2-interacting lncRNA NRON, which promotes tumor formation by suppressing both P53-dependent and independent pathways. NRON binds to MDM2 and MDMX (MDM4) via two different stem-loops respectively and induces their heterogenous dimerization, thereby enhancing the E3 ligase activity of MDM2 towards its tumor suppressing substrates, including P53, RBI and NFATI, etc. Overall design: To identify those lncRNAs that interact with MDM2 and function in both P53-dependent and -independent ways, MDA-MB-231 and MCF-7 cells with exogenous Flag-tagged MDM2 were established and subjected to RNA immunoprecipitation (RIP) using negative control normal mouse IgG and anti-FLAG antibody respectively. RIP–sequencing (RIP-seq) was then performed to identify the lncRNAs that specifically bind to FLAG-tagged MDM2 but not to normal mouse IgG control.