DDX3X functionally and physically interacts with Estrogen Receptor-alpha

DEAD-box protein 3X (DDX3X) is a human DEAD-box protein with conventional roles in RNA metabolism and unconventional functions in signalling pathways that do not require its enzymatic activity. For example, DDX3X acts as a multifunctional adaptor molecule in anti-viral innate immune signalling pathways, where it interacts with and regulates the kinase IKB-kinase-epsilon (IIKK epsilon). Interestingly, both DDX3X and IKK epsilon have also independently been shown to act as breast cancer oncogenes. IKK epsilon's oncogenic functions are likely multifactorial, but it was suggested to phosphorylate the transcription factor Estrogen receptor alpha (ER alpha) at Serine 167, which drives expression of Er alpha target genes in an estrogen-independent manner. In this study, we identified a novel physical interaction between DDX3X and ER alpha that positively regulates ER alpha activation. DDX3X knockdown in ER+ breast cancer cell lines resulted in reduced ER alpha phosphorylation, reduced Estrogen Response Element (ERE)-controlled reporter gene expression, decreased expression of ER alpha target genes, and decreased cell proliferation. Vice versa, overexpression of DDX3X resulted in enhanced ER alpha phosphorylation and activity. Furthermore, we provide evidence that DDX3X physically binds to ER alpha from co-immunoprecipitation and pulldown experiments. Based on our data, we propose that DDX3X acts as an adaptor to facilitate IKK epsilon-mediated ER alpha activation, akin to the mechanism we previously elucidated for IKK epsilon-mediated Interferon Regulatory factor 3 (IRF3) activation in innate immune signalling. In conclusion, our research provides a novel molecular mechanism that might contribute to the oncogenic effect of DDX3X in breast cancer, potentially linking it to the development of resistance against endocrine therapy.