Murine double minute 2 regulates Hu antigen R stability in human liver and colon cancer through NEDDylation

HuR, a member of the Elav/Hu family, is a gene expression regulator with a central role in mRNA stabilization (1), HuR has been shown to increase cellular division by enhancing the stability of mRNAs encoding important proteins for cell cycle control and proliferation, as well as other factors that influence tumor cell growth (2, 3). HuR expression is elevated in colon, breast, prostate, pancreatic, oral, skin, brain, gastric, lung and ovarian cancers, and correlates with tumor malignancy, supporting a role for HuR in tumorigenesis (4–6). In the liver, HuR plays a crucial role in hepatocyte proliferation, differentiation, and HCC transformation (7). At the transcriptional level, NF-κB can activate HuR transcription downstream of the PI3K-AKT signaling pathway (8). At the post-transcriptional level, the ubiquitin (Ub)-proteasome pathway has been implicated in HuR function, whereas the machinery involved in enhancing HuR protein stability remains obscure (9). The ubiquitin-like molecule NEDD8 is a key regulator of cell growth, viability and development. NEDD8 is ubiquitously expressed and highly conserved among most eukaryotes. At the molecular level, only members of the cullin family of proteins, are well-characterized substrates for NEDDylation (10). However, through direct biological approaches or proteomic techniques, it is evident that NEDD8 proteome is widely diverse. Recent studies have shown that ribonucleoproteins are targets for NEDD8 conjugation which protect them from de-stabilization (11, 12). Conversely, cysteine protease (NEDP1) removes NEDD8 molecules from conjugated substrates (13). Mdm2 acts as an E3 NEDD8 ligase, promoting p53 NEDDylation and stabilization (14), and, of importance, Mdm2 is overexpressed in many human tumors, at least in part due to gene amplification (15). In this study, we show that HuR is overexpressed in proliferative HCC and colon cancer cells and biopsies through Mdm2-mediated NEDDylation. Our data suggests that HuR NEDDylation at K313 and K326 residues stabilize HuR. Mechanistically, Mdm2-mediated NEDDylation of HuR controls the nuclear localization of HuR and protects it from degradation. To our knowledge, this is the first report to implicate NEDDylation in the regulation of HuR stability and localization. Our findings show that NEDDylation is a novel mechanism for HuR regulation that might represent a useful tool for new therapeutic strategies for HCC and colon cancer.