Myosin regulation of NKCC1: effects on cAMP-mediated Cl- secretion in intestinal epithelia.
The basally located actin cytoskeleton has been demonstrated previously to regulate Cl- secretion from intestinal epithelia via its effects on the Na+-K+-2Cl- cotransporter (NKCC1). In nontransporting epithelia, inhibition of myosin light chain kinase (MLCK) prevents cell-shrinkage-induced activation of NKCC1. The aim of this study was to investigate the role of myosin in the regulation of secretagogue-stimulated Cl- secretion in intestinal epithelia. The human intestinal epithelial cell line T84 was used for these studies. Prevention of myosin light chain phosphorylation with the MLCK inhibitor ML-9 or ML-7 and inhibition of myosin ATPase with butanedione monoxime (BDM) attenuated cAMP but not Ca2+-mediated Cl- secretion. Both ML-9 and BDM diminished cAMP activation of NKCC1. Neither apical Cl- channel activity, basolateral K+ channel activity, nor Na+-K+-ATPase were affected by these agents. Cytochalasin D prevented such attenuation. cAMP-induced rearrangement of basal actin microfilaments was prevented by both ML-9 and BDM. The phosphorylation of mosin light chain and subsequent contraction of basal actin-myosin bundles are crucial to the cAMP-driven activation of NKCC1 and subsequent apical Cl- efflux.
Hecht
G
G
Section of Digestive and Liver Diseases, Department of Medicine, University of Illinois and West Side Veterans Affairs Medical Center, Chicago, Illinois 60612, USA. gahecht@uic.edu
Koutsouris
A
A
D000199:Actins / Q000502:physiology; D001381:Azepines / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018118:Chloride Channels / Q000187:drug effects; D002712:Chlorides / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology*; D003599:Cytoskeleton / Q000502:physiology; D003931:Diacetyl / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D018994:Myosin Light Chains / Q000037:antagonists & inhibitors / Q000378:metabolism; D009218:Myosins / Q000037:antagonists & inhibitors / Q000502:physiology*; D010766:Phosphorylation; D015221:Potassium Channels / Q000187:drug effects; D028021:Sodium-Potassium-Chloride Symporters
D000199:Actins; D001381:Azepines; D002352:Carrier Proteins; D018118:Chloride Channels; D002712:Chlorides; D018994:Myosin Light Chains; D015221:Potassium Channels; D028021:Sodium-Potassium-Chloride Symporters; C056218:ML 9; C004717:diacetylmonoxime; D000242:Cyclic AMP; D009218:Myosins; D003931:Diacetyl