aPKC kinase activity is required for the asymmetric differentiation of the premature junctional complex during epithelial cell polarization.
We have previously shown that aPKC interacts with cell polarity proteins PAR-3 and PAR-6 and plays an indispensable role in cell polarization in the C. elegans one-cell embryo as well as in mammalian epithelial cells. Here, to clarify the molecular basis underlying this aPKC function in mammalian epithelial cells, we analyzed the localization of aPKC and PAR-3 during the cell repolarization process accompanied by wound healing of MTD1-A epithelial cells. Immunofluorescence analysis revealed that PAR-3 and aPKClambda translocate to cell-cell contact regions later than the formation of the primordial spot-like adherens junctions (AJs) containing E-cadherin and ZO-1. Comparison with three tight junction (TJ) membrane proteins, JAM, occludin and claudin-1, further indicates that aPKClambda is one of the last TJ components to be recruited. Consistently, the expression of a dominant-negative mutant of aPKClambda (aPKClambdakn) in wound healing cells does not inhibit the formation of the spot-like AJs; rather, it blocks their development into belt-like AJs. These persistent spot-like AJs in aPKClambda-expressing cells contain all TJ membrane proteins and PAR-3, indicating that aPKC kinase activity is not required for their translocation to these premature junctional complexes but is indispensable for their further differentiation into belt-like AJs and TJs. Cortical bundle formation is also blocked at the intermediate step where fine actin bundles emanating from premature cortical bundles link the persistent spot-like AJs at apical tips of columnar cells. These results suggest that aPKC contributes to the establishment of epithelial cell polarity by promoting the transition of fibroblastic junctional structures into epithelia-specific asymmetric ones.
Department of Molecular Biology, Yokohama City University School of Medicine, 3-9 Fuku-ura, Kanazawa-ku, Yokohama 236-0004, Japan.
Atsushi
A
Suzuki
Chikako
C
Ishiyama
Katsutaka
K
Hashiba
Miki
M
Shimizu
Klaus
K
Ebnet
Shigeo
S
Ohno
D008841:Actin Cytoskeleton / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002450:Cell Communication / Q000502:physiology; D018797:Cell Cycle Proteins; D002454:Cell Differentiation / Q000502:physiology*; D016764:Cell Polarity / Q000502:physiology*; D002478:Cells, Cultured; D062445:Claudin-1; D004847:Epithelial Cells / Q000166:cytology / Q000201:enzymology*; D005455:Fluorescent Antibody Technique; D007365:Intercellular Junctions / Q000201:enzymology* / Q000648:ultrastructure; D007527:Isoenzymes; D062726:Junctional Adhesion Molecules; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D009154:Mutation / Q000502:physiology; D062793:Occludin; D010750:Phosphoproteins / Q000378:metabolism; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D014945:Wound Healing / Q000502:physiology; D062826:Zonula Occludens-1 Protein; D000099293:Protein Kinase C-lambda
D048868:Adaptor Proteins, Signal Transducing; D015820:Cadherins; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D018797:Cell Cycle Proteins; D062445:Claudin-1; D007527:Isoenzymes; D062726:Junctional Adhesion Molecules; D008565:Membrane Proteins; D062793:Occludin; D010750:Phosphoproteins; D011493:Protein Kinase C; D062826:Zonula Occludens-1 Protein; D000099293:Protein Kinase C-lambda; C568113:Cldn1 protein, mouse; C568986:Ocln protein, mouse; C115084:Pard3 protein, mouse; C569043:Tjp1 protein, mouse; C115015:PKC-3 protein