Dysregulation of ZnT2‐Mediated Zn Homeostasis Impairs Epithelial Polarity And Breast Differentiation

Abstract

Epithelial polarity is essential for the architecture and function of the mammary gland. We previously found that the loss of the vesicular zinc (Zn) transporter ZnT2 in ZnT2‐null mice compromises polarity, reduces STAT5 activation and impairs mammary gland differentiation. Here, we aimed to determine how the loss of ZnT2 impairs polarity and mammary gland differentiation. We used ZnT2‐attenuated mammary epithelial cells (MECs) in vitro and hypothesized that the loss of ZnT2‐mediated Zn transport dysregulates (PTEN), an epithelial polarity factor that disrupts spatial orientation of polarity, which subsequently impairs prolactin‐mediated Jak2/STAT5 signaling. First, we found that without the ability to sequester Zn into vesicles via ZnT2, Zn accumulated in the cytoplasm and significantly reduced PTEN expression. Moreover, ZnT2‐attenuated MECs had reduced membrane expression of, Annexin A2, a protein involved in apical membrane transport, and disrupted localization of ZO‐1, a protein critical for tight junction assembly, indicating impaired spatial orientation. Consequently, ZnT2‐attenuated MECs failed to differentiate into 3D mammospheres with expanded alveolar lumen. Importantly, we showed that the loss polarity disrupted cell surface localization of the principal lactogenic hormone receptor, prolactin receptor, and reduced downstream Jak2/STAT5 signaling, collectively indicating that cytoplasmic Zn accumulation dysregulates epithelial polarity and mammary differentiation. Our recent studies in breastfeeding women identified several non‐synonymous genetic variants in ZnT2 that result in “loss‐of‐function”. To investigate effects of these variants (ZnT2K66N and ZnT2Q71H) on MEC polarity, we transfected MECs to express these ZnT2 variants and found that expression increased cytoplasmic Zn and reduced PTEN expression in MECs, implicating defects of ZnT2 function in breast differentiation in women. Taken together, these results establish that ZnT2‐mediated Zn homeostasis is critical for maintaining epithelial polarity and regulating prolactin signaling for proper mammary gland differentiation and this suggests that “loss‐of‐function” variants of ZnT2 may be associated with impaired breast differentiation and lactation insufficiency in women.Support or Funding InformationIntramural funds to SLK