Myosin light chain phosphorylation regulates barrier function by remodeling tight junction structure.
Epithelial tight junctions form a barrier against passive paracellular flux. This barrier is regulated by complex physiologic and pathophysiologic signals that acutely fine-tune tight junction permeability. Although actomyosin contraction and myosin light chain phosphorylation are clearly involved in some forms of tight junction regulation, the contributions of other signaling events and the role of myosin light chain phosphorylation in this response are poorly understood. Here we ask if activation of myosin light chain kinase alone is sufficient to induce downstream tight junction regulation. We use a confluent polarized intestinal epithelial cell model system in which constitutively active myosin light chain kinase, tMLCK, is expressed using an inducible promoter. tMLCK expression increases myosin light chain phosphorylation, reorganizes perijunctional F-actin, and increases tight junction permeability. TJ proteins ZO-1 and occludin are markedly redistributed, morphologically and biochemically, but effects on claudin-1 and claudin-2 are limited. tMLCK inhibition prevents changes in barrier function and tight junction organization induced by tMLCK expression, suggesting that these events both require myosin light chain phosphorylation. We conclude that myosin light chain phosphorylation alone is sufficient to induce tight junction regulation and provide new insights into the molecular mechanisms that mediate this regulation.
Shen
Le
L
Department of Pathology, The University of Chicago, 5841 South Maryland Avenue, MC 1089,Chicago, IL 60637, USA.
Black
Eric D
ED
Witkowski
Edwina D
ED
Lencer
Wayne I
WI
Guerriero
Vince
V
Schneeberger
Eveline E
EE
Turner
Jerrold R
JR
D000199:Actins / Q000378:metabolism; D018938:Caco-2 Cells; D002463:Cell Membrane Permeability / Q000502:physiology; D062445:Claudin-1; D057167:Claudins; D004790:Enzyme Induction; D004848:Epithelium / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D018994:Myosin Light Chains / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000096:biosynthesis / Q000378:metabolism*; D062793:Occludin; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D019108:Tight Junctions / Q000201:enzymology / Q000378:metabolism* / Q000648:ultrastructure; D062826:Zonula Occludens-1 Protein
D000199:Actins; C568108:CLDN1 protein, human; C491875:CLDN2 protein, human; D062445:Claudin-1; D057167:Claudins; D008565:Membrane Proteins; D018994:Myosin Light Chains; C568985:OCLN protein, human; D062793:Occludin; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D009219:Myosin-Light-Chain Kinase