ZnR/GPR39, BY REGULATING KCC ACTIVITY, ENHANCES ESTROGEN NEGATIVE BREAST CANCER CELLS GROWTH

Breast cancer malignant cells exhibit deviant signaling patterns and changes in Zn²⁺ accumulation and distribution but the link between these pathways is not well understood. Our study shows that extracellular Zn²⁺ triggers ZnR/GPR39, a Gq-coupled receptor, linked to cell growth. KCC cotransporters mediate K⁺/ Cl^- transport across the plasma membrane and have been implied to play a role in cell migration and proliferation. We hypothesized that Zn²⁺ via its sensing receptor ZnR/GPR39 stimulates KCC and leads to breast cancer cell growth. Zn²⁺-dependent Ca²⁺ response, mediated by ZnR/GPR39, was measured in estrogen negative BT20 cells, while the effect was inhibited in cells silenced with siGPR39 construct. Transport activity by KCC was enhanced by ZnR/GPR39 activation, monitored as the rate of NH₄⁺ transport used as a surrogate to K⁺, and this was attenuated by silencing of GPR39. Similarly, inhibitors of KCC (DIOA), Gq (YM254890) or mitogen activated protein kinase (MAPK, UO126) abolished the effect of Zn²⁺. Our results therefore propose that Zn²⁺ via ZnR/GPR39 activates MAPK pathway and induces KCC-dependent ion transport in breast cancer cells. Moreover, a scratch assay was performed on BT20 confluent cultures to determine if this pathway enhances cell proliferation and migration. Activation of ZnR/GPR39, with Zn²⁺, enhances scratch closure rates and this is attenuated by inhibition of KCC (using DIOA) or silencing of ZnR/GPR39. Hence, we suggest that in estrogen negative BT20 breast cancer cells ZnR/GPR39 regulates KCC-dependent ion transport and cell growth and may therefore contribute to breast cancer progression.