Background: The primary cilium is a microtubule-based and nonmotile organelle functioning as a cellular antenna that is involved in the regulation of cell proliferation, differentiation, and migration. In breast cancer cells, the primary cilium is a structure that decreases in incidence with increasing degrees of transformation and may be biologically more important in estrogen receptor (ERα)-negative breast cancer cells. Split ends (SPEN) is an ERα corepressor that we have identified as a tumor suppressor protein in ERα-positive breast cancer cells whose hormone-independent roles in breast cancer have never been explored.
Methods: We determined the hormone-independent transcriptional program regulated by the ERα cofactor SPEN in breast cancer using DNA microarrays. The biological functions regulated by SPEN independently of hormones were studied in vitro in ERα-positive and ERα-negative breast cancer cells. Finally, we examined the clinical relevance of SPEN expression in cohorts of breast cancer samples with outcome data.
Results: We found that SPEN is coexpressed with a number of genes involved in ciliary biology, including the ciliogenic transcription factor RFX3, in a hormone-independent manner. SPEN reexpression in T47D cells containing a nonsense mutation in SPEN restored the primary cilium, whereas its knockdown in MCF10A and Hs578T cells considerably decreased primary cilia levels. We also report that SPEN regulates migration in breast cells, but only in those harboring primary cilia, and that KIF3A silencing, a critical factor in primary cilia, partially reverses SPEN's effects, suggesting that SPEN may coordinate cellular movement through primary cilia-dependent mechanisms. Finally, we found that high SPEN RNA levels were predictive of early metastasis in two independent cohorts of 77 (HR 2.25, P = 0.03) and 170 (HR = 2.23, P = 0.004) patients with ERα-negative breast cancer.
Conclusions: Together, our data demonstrate a role for SPEN in the regulation of primary cilia formation and cell migration in breast cancer cells, which may collectively explain why its expression is associated with time to metastasis in cohorts of patients with ERα-negative breast cancers.
Keywords: Breast cancer; Cell migration; Metastasis; Primary cilia.