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  • Kcnj10 is a major type of K+ channel in mouse corneal epithelial cells and plays a role in initiating EGFR signaling.

Kcnj10 is a major type of K+ channel in mouse corneal epithelial cells and plays a role in initiating EGFR signaling.

American journal of physiology. Cell physiology (2014-08-08)
Lijun Wang, Chengbiao Zhang, Xiaotong Su, Daohong Lin
ABSTRACT

We used primary mouse corneal epithelial cells (pMCE) to examine the role of Kcnj10 in determining membrane K(+) conductance and cell membrane potential and in regulating EGF/TGFA release. Western blot, immunostaining, and RT-PCR detected the expression of Kcnj10 in mouse cornea. The single channel recording identified the 20-pS inwardly rectifying K(+) channels in pMCE of WT mice, but these channels were absent in Kcnj10(-/-). Moreover, the whole cell recording demonstrates that deletion of Kcnj10 largely abolished the inward K(+) currents and depolarized the cell membrane K(+) reversal potential (an index of the cell membrane potential). This suggests that Kcnj10 is a main contributor to the cell K(+) conductance and it is pivotal in generating membrane potential in cornea. Furthermore, to test the hypothesis that Kcnj10 expression plays a key role in the stimulation of growth factors release, we employed an immortalized human corneal epithelial cell line (HCE) transfected with siRNA-Kcnj10 or siRNA-control. Levels of TGFA and EGF secreted in the medium were measured by ELISA. Coimmunoprecipitation, biotinylation, and pull-down assay were used to examine the expression of EGFR and the GTP bound form of Rac1 (active Rac1). Downregulation of Kcnj10 activated Rac1 and enhanced EGF/TGFA release, which further contributed to the upregulation of EGFR phosphorylation and surface expression. We conclude that Kcnj10 is a main K(+) channel expressed in corneal epithelial cells and the inhibition of Kcnj10 resulted in depolarization, which in turn induced an EGF-like effect.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Mouse EGF ELISA Kit, for serum, plasma and cell culture supernatant
Sigma-Aldrich
Rat Egf / Pro-epidermal Growth Factor ELISA Kit, for serum, plasma and cell culture supernatants
Sigma-Aldrich
Human EGF ELISA Kit, for serum, plasma, cell culture supernatant.