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Gender specific inhibition of KCNQ1 in the antisecretory effect of 17β‐estradiol on rat distal colonic crypts is regulated by PKCδ
Author(s) -
O’Mahony Fiona,
Alzamora Rodrigo,
Betts Vicki,
Harvey Brian J
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a799-b
Subject(s) - rottlerin , protein kinase c , ussing chamber , secretion , endocrinology , medicine , intracellular , epithelial sodium channel , cystic fibrosis transmembrane conductance regulator , chemistry , biology , microbiology and biotechnology , signal transduction , sodium , organic chemistry , cystic fibrosis
Our previous work has shown a female gender specific inhibitory effect of 17β‐estradiol (E2) on intestinal Cl ‐ secretion, involving inhibition of basolateral K + conductance. This study investigated KCNQ1 as a candidate K + channel in the E2 anti‐secretory effect. Immunoblot and RT‐PCR analysis of male and female isolated colonic crypts demonstrated similar expression of KCNQ1. Colonic epithelia were mounted in Ussing chambers and short circuit current ( I SC ) was measured as an index of Cl − secretion, stimulated using heat‐stable enterotoxin (STa, 50 nM). E2 (1–100nM) decreased the STa‐stimulated I SC within 5 minutes in a dose dependent manner (1nM E2, 85±7%; 10nM, 68±5%; 100 nM, 47±4%). The E2 effect was sensitive to the PKC™ inhibitor Rottlerin (10 μM). E2 (10 nM) inhibited a chromanol 293B‐sensitive basolateral K + conductance in nystatin‐perforated epithelia (control 57±6, E2 7±3 μA/cm 2 ), indicating KCNQ1 involvement. Immunoblot and RT‐PCR analysis showed a higher basal expression of PKCδ in female crypts compared to male. No difference was observed in PKA levels. E2 10 nM induced activation of PKCδ (2 min) and PKA (2 min) only in female. PKA activity was inhibited by Rottlerin, indicating downstream activation from PKCδ . E2 treatment induced translocation of PKCδ to the plasma membrane and association to KCNQ1 in female crypts. In Vitro studies using GST‐KCNQ1 intracellular termini showed that PKCδ and PKA phosphorylate the N‐termini. These data suggests that PKCδ may act as key regulator of female specific E2 targeting of KCNQ1 in the inhibition of Cl − secretion. Supported by HEA‐PRTLI Programme.