Voltage-gated Potassium Channel 1.5 And Activation Of MAPK Signaling Pathway Induced By Cav-1 In Human Mammary Epithelial Cells

Potassium channels (K~+ channel) are membrane-spanning proteins that selectively conduct K ions across the cell membrane along its electrochemical gradient, and are involved in many physiological functions, such as proliferation, solute transport, volume control, enzyme activity, secretion, invasion, gene expression, excitation-contraction coupling and intercellular communication. Recent reports indicate that Caveolae, a special lipid raft in cell membrane, not only gathers signaling molecules, but also distributes a number of ion channels. Moreover, it also works together with ion channels to regulate the intracellular ion transport and signal transduction. Caveolin-1 (Cav-1) is a principal component of Caveolae membranes that function as scaffold proteins for multiple membrane-initiated signaling pathways. Cav-1 can participate in a variety of intracellular activities, such as vesicular transport, cellular cholesterol homeostasis, signal transduction, oncogenes and tumorigenesis. Our lab has reported that blocking Kv channels can significantly inhibit cell proliferation in human breast cancer cell (MCF7) and Cav-1 haploinsufficient cell (MCF10A-ST1). What's more, Kv1.5 had related with Cav-1. However, their precise mechanisms are still unknown. In this study, we researched the relationship between potassium channels properties, expression and the activation of MAPK signaling pathway which could induced by Cav-1 using Patch clamp and Western blot technology. Meanwhile, in order to investigate the role of Kv1.5 in the relationship between Cav-1 and MAPK signaling pathway, we transfected a plasmid named pRBG4-Kv1.5 which could overexpress Kv1.5 in MCF7 cells to understand the relationship between Kv channels and Cav-1 which could affect the proliferation of breast cell , the mechanism of signal transductions. Results showed that:(1) There was a voltage-dependent, outward rectification K~+ currents in MCF10A cells which could inhibit by 4-AP, and the current was different in MCF10A and MCF10A-ST1 cells. It provided that, down regulation of Cav-1 might affect types and expression of Kv channels in MCF10A and MCF10A-ST1 cells.(2) Using MβCD in MCF10A cells can decrease the number of Caveolae and downregulate the expression of Kv1.1, Kv1.2, Kv1.3 and Kv1.5 mRNA, the change of Kv1.5 was the most significant.(3) After transiently transfection of pRBG4-Kv1.5 plasmid for 72h in MCF7 cell, the expression of Cav-1 protein upregulated, further research indicated that the phosphorylation of ERK1/2 increased, and MAPK signal pathway was activated. (4) Inhibiting Kv channels in MCF10A and MCF10A-ST1 cells by 4-AP, the sensitivity of 4-AP is different in breast cells which Cav-1 expression is different and the level of MAPK activation is different.Conclusion: Voltage-gated potassium channel 1.5 had participated in the regulation of MAPK signaling transduction in mammary epithelial cells, and was related with the Cav-1.