Cytoplasmic Localization of Proline, Glutamic Acid, Leucine-rich Protein 1 (PELP1) Induces Breast Epithelial Cell Migration through Up-regulation of Inhibitor of κB Kinase ϵ and Inflammatory Cross-talk with Macrophages

Abstract Cytoplasmic localization of proline, glutamic acid, leucine rich protein 1 (PELP1) is observed in about 40% of women with invasive breast cancer. In mouse models, PELP1 overexpression in the mammary gland leads to pre-malignant lesions and eventually mammary tumors. In preliminary clinical studies, cytoplasmic localization of PELP1 was seen in 36% of women at high risk of developing breast cancer. Here, we investigated whether cytoplasmic PELP1 signaling promotes breast cancer initiation in models of immortalized human mammary epithelial cells (HMECs). Global gene expression (GGE) analysis was performed on HMEC lines expressing vector control, PELP1-wt, or mutant PELP1 in which the nuclear localization sequence was altered, resulting in cytoplasmic localization of PELP1 (PELP1-cyto). GGE analysis identified that PELP1-cyto expression in HMECs induced NF-κB signaling pathways. Western blot analysis of PELP1-cyto HMECs showed upregulation of inhibitor of kappa B kinase epsilon (IKKe) and increased phosphorylation of the NF-κB subunit RelB. To determine whether secreted factors produced by PELP1-cyto HMECs promote macrophage activation, THP-1 macrophages were treated with HMEC-conditioned media (CM). PELP1-cyto CM induced changes in THP-1 gene expression as compared to control cell CM. Double conditioned media (DCM) from the activated THP-1 cells was then applied to HMECs to determine whether paracrine signaling from PELP1-cyto-activated macrophages, could in turn, promote migration of HMECs. PELP1-cyto DCM induced robust HMEC migration, which was reduced in DCM from PELP1-cyto HMECs expressing IKKe shRNA. Our findings suggest cytoplasmic localization of PELP1 upregulates pro-tumorigenic IKKe and secreted inflammatory signals, which through paracrine macrophage activation, regulate the migratory phenotype associated with breast cancer initiation.