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.
Ivanov
Andrei I
AI
Department of Medicine, University of Rochester, Rochester, NY 14642, USA. Andrei_Ivanov@urmc.rochester.edu
Samarin
Stanislav N
SN
Bachar
Moshe
M
Parkos
Charles A
CA
Nusrat
Asma
A
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