Expression of the catalytic domain of myosin light chain kinase
increases paracellular permeability.
Hecht, Gail, Lidija Pestic, Gordana Nikcevic, Athanasia Koutsouris,
Jyoti Tripuraneni, Donald D. Lorimer, Grzegorz Nowak, Vince
Guerriero, Jr., Elliot L. Elson, and Primal De Lanerolle.
Department of Medicine, Department of Physiology and Biophysics,
University of Illinois at Chicago, Chicago, IL 60612, Department of
Animal Sciences, University of Arizona, Tucson, AR and Department of
Biochemistry and Molecular Biophysics, Washington University Medical
School, St. Louis, MO
APStracts 3:0166C, 1996.
Contractile events resulting from myosin light chain (MLC20)
phosphorylation have been implicated in the regulation of epithelial
tight junction permeability. To address this question,Madin- Darby
canine kidney (MDCK) cells were transfected with a murine leukemia
retroviral vector containing DNA encoding either the catalytic domain
of myosin light chain kinase (tMK) or the [beta]-galactosidase gene
([beta]-gal). Autoradiograms of SDS-PAGE analysis of myosin
immunoprecipitated from 32Pi-labelled transfected cells demonstrated
that MLC20 phosphorylation was increased 3.1+0.9-fold in cells
expressing tMK compared to cells expressing [beta]-gal.
Phosphopeptide mapping confirmed that myosin light chain kinase
(MLCK) was responsible for the increased MLC20 phosphorylation.
Transepithelial electrical resistance, a measurement of barrier
function, of tMK cell monolayers was consistently less than 10% (123
+ 20 [omega].cm2) of that of monolayers comprised of wild type cells
(1456 + 178 [omega].cm2) or cells expressing [beta]-gal (1452 + 174
[omega].cm2). Dual 22Na+ and 3H-mannitol flux studies indicated that
the decrease in resistance in tMK cells was attributable to increased
paracellular flow. These data support the idea that MLC20
phosphorylation by MLCK is involved in regulating epithelial tight
junction permeability.
Received 19 January 1996; accepted in final form 9 May 1996.
APS Manuscript Number C30-6.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 5 June 96