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 (IIKKε). Interestingly, both DDX3X and IKKɛ have also independently been shown to act as breast cancer oncogenes. IKKɛ's oncogenic functions are likely multifactorial, but it was suggested to phosphorylate the transcription factor Estrogen receptor alpha (ERα) at Serine 167, which drives expression of Erα target genes in an estrogen-independent manner. In this study, we identified a novel physical interaction between DDX3X and ERα that positively regulates ERα activation. DDX3X knockdown in ER+ breast cancer cell lines resulted in reduced ERα phosphorylation, reduced Estrogen Response Element (ERE)-controlled reporter gene expression, decreased expression of ERα target genes, and decreased cell proliferation. Vice versa, overexpression of DDX3X resulted in enhanced ERα phosphorylation and activity. Furthermore, we provide evidence that DDX3X physically binds to ERα from co-immunoprecipitation and pulldown experiments. Based on our data, we propose that DDX3X acts as an adaptor to facilitate IKKε-mediated ERα activation, akin to the mechanism we previously elucidated for IKKε-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.