Objective:It has been shown that LRP16 is an estrogen-induced gene through its receptor α(ERα).Although there is evidence demonstrating that inhibition of LRP16 gene expression in MCF-7 human breast cancer cells partially attenuates its estrogen-responsiveness,the underlying molecular mechanism is still unclear.Here,the effect of LRP16 expression on the ERα signaling transduction was investigated.Methods:Cotransfection assays were used to measure the effect of LRP16 on ERα-mediated transcriptional activity.GST-pulldown and immunoprecipitation(CoIP)assays were employed to investigate the physical interaction of LRP16 and ERα.The mammalian two-hybrid method was used to map the functional interaction region.Results:the results of cotransfection assays demonstrated that the transcriptional activities of ERα were enhanced in a LRP16 dose-dependent manner in MCF-7 in the presence of estrogen,however,it was abolished in the absence of E2 in MCF-7 cells.The physical interaction of LRP16 and ERα proteins was confirmed by GST-pulldown in vitro and CoIP in vivo assays,which was enhanced by E2 but not dependent on its presence.Furthermore,the results of the mammalian two-hybrid assays indicated that the binding region of ERα to LRP16 located at the A/B AF-1 functional domain and E2 stimulated the binding of LRP16 to the full-length ERα molecule but not to the A/B region alone.Conclusion:These results support a role for estrogenically regulated LRP16 as an ERα coactivator,providing a positive feedback regulatory loop for ERα signal transduction.Based on this function of LRP16,we propose that ERα-positive breast cancer patients with high expression of LRP16 might benefit from targeting LRP16 therapy.
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