CK1d modulates the transcriptional activity of ERa via AIB1 in an estrogen-dependent manner and regulates ERa–AIB1 interactions

Oncogenesis in breast cancer often requires the overexpression of the nuclear receptor coactivator AIB1/SRC-3 acting in conjunction with estrogen receptor-a (ERa). Phosphorylation of both ERa and AIB1 has been shown to have profound effects on their functions. In addition, proteasome-mediated degradation plays a major role by regulating their stability and activity. CK1d, a member of the ubiquitous casein kinase-1 family, is implicated in the progression of breast cancer. In this study, we show that both ERa and AIB1 are substrates for CK1d in vitro, and identify a novel AIB1 phosphorylation site (S601) targeted by CK1d, significant for the co-activator function of AIB1. CK1d is able to interact with ERa and AIB1 in vivo, while overexpression of CK1d in breast cancer cells results in an increased association of ERa with AIB1 as confirmed by co-immunoprecipitation assays from cell lysates. Using an siRNA-based approach, luciferase reporter assays and qRT-PCR, we observe that silencing of CK1d leads to reduced ERa transcriptional activity, despite increased ERa levels, similarly to proteasome inhibition. We provide evidence that AIB1 protein levels are reduced by CK1d silencing, in an estradiol-dependent manner; such destabilization can be inhibited by pre-treatment with the proteasome inhibitor MG132. We propose that differing activities adopted by ERa and AIB1 as a consequence of their interactions with and phosphorylation by CK1d, particularly AIB1 stabilization, influence the transcriptional activity of ERa, and therefore have a role in breast cancer development.