Identification of interacting proteins of retinoid-related orphan nuclear receptor gamma in HepG2 cells.

ABSTRACT

The retinoid-related orphan nuclear receptor gamma (ROR γ) plays critical roles in regulation of development, immunity and metabolism. As transcription factor usually forms a protein complex to function, thus capturing and dissecting of the ROR γ protein complex will be helpful for exploring the mechanisms underlying those functions. After construction of the recombinant tandem affinity purification (TAP) plasmid, pMSCVpuro ROR γ-CTAP(SG), the nuclear localization of ROR γ-CTAP(SG) fusion protein was verified. Following isolation of ROR γ protein complex by TAP strategy, seven candidate interacting proteins were identified. Finally, the heat shock protein 90 (HSP90) and receptor-interacting protein 140 (RIP140) were confirmed to interplay with ROR γ by co-immunoprecipitation. Interference of HSP90 or/and RIP140 genes resulted in dramatically decreased expression of CYP2C8 gene, the ROR γ target gene. Data from this study demonstrate that HSP90 and RIP140 proteins interact with ROR γ protein in a complex format and function as co-activators in the ROR γ-mediated regulatory processes of HepG2 cells.