title: Kcnj10 is a major type of K+ channel in mouse corneal epithelial cells and plays a role in initiating EGFR signaling.
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.
authors.affiliation: Department of Pharmacology, New York Medical College, Valhalla, New York.
authors.forename: Lijun
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, New York Medical College, Valhalla, New York.
authors.forename: Chengbiao
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology, New York Medical College, Valhalla, New York.
authors.forename: Xiaotong
authors.initials: X
authors.lastname: Su
authors.affiliation: Department of Pharmacology, New York Medical College, Valhalla, New York daohong_lin@nymc.edu.
authors.forename: Daohong
authors.initials: D
authors.lastname: Lin
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004815:Epidermal Growth Factor / Q000378:metabolism; D019573:Epithelium, Corneal / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D008564:Membrane Potentials; D018345:Mice, Knockout; D011188:Potassium / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D016211:Transforming Growth Factor alpha / Q000378:metabolism; D000098854:Kcnj10 Channel
chemical_list: D000098854:Kcnj10 Channel; D024661:Potassium Channels, Inwardly Rectifying; D016211:Transforming Growth Factor alpha; D004815:Epidermal Growth Factor; C541485:EGFR protein, mouse; D066246:ErbB Receptors; D011188:Potassium
keywords: EGFR signaling; Kir4.1; Rac1; corneal epithelial cells membrane potential
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