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. 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.