Protein kinase C activation disrupts epithelial apical junctions via ROCK-II dependent stimulation of actomyosin contractility.
BACKGROUND
Disruption of epithelial cell-cell adhesions represents an early and important stage in tumor metastasis. This process can be modeled in vitro by exposing cells to chemical tumor promoters, phorbol esters and octylindolactam-V (OI-V), known to activate protein kinase C (PKC). However, molecular events mediating PKC-dependent disruption of epithelial cell-cell contact remain poorly understood. In the present study we investigate mechanisms by which PKC activation induces disassembly of tight junctions (TJs) and adherens junctions (AJs) in a model pancreatic epithelium.


RESULTS
Exposure of HPAF-II human pancreatic adenocarcinoma cell monolayers to either OI-V or 12-O-tetradecanoylphorbol-13-acetate caused rapid disruption and internalization of AJs and TJs. Activity of classical PKC isoenzymes was responsible for the loss of cell-cell contacts which was accompanied by cell rounding, phosphorylation and relocalization of the F-actin motor nonmuscle myosin (NM) II. The OI-V-induced disruption of AJs and TJs was prevented by either pharmacological inhibition of NM II with blebbistatin or by siRNA-mediated downregulation of NM IIA. Furthermore, AJ/TJ disassembly was attenuated by inhibition of Rho-associated kinase (ROCK) II, but was insensitive to blockage of MLCK, calmodulin, ERK1/2, caspases and RhoA GTPase.


CONCLUSIONS
Our data suggest that stimulation of PKC disrupts epithelial apical junctions via ROCK-II dependent activation of NM II, which increases contractility of perijunctional actin filaments. This mechanism is likely to be important for cancer cell dissociation and tumor metastasis.
Department of Medicine, University of Rochester, Rochester, NY 14642, USA. Andrei_Ivanov@urmc.rochester.edu
Andrei I
AI
Ivanov
Stanislav N
SN
Samarin
Moshe
M
Bachar
Charles A
CA
Parkos
Asma
A
Nusrat
D000205:Actomyosin / Q000235:genetics / Q000502:physiology*; D022005:Adherens Junctions / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D024462:Myosin Type II / Q000378:metabolism; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D019108:Tight Junctions / Q000235:genetics / Q000378:metabolism*; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism*
D000205:Actomyosin; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D024462:Myosin Type II