HSP27 modulates agonist-induced association of translocated RhoA and PKC-alpha in muscle cells of the colon.
The recruitment of signal transduction molecules to the membrane is crucial for the efficient coupling of extracellular signals and contractile response. The trafficking is dynamic. We have investigated a possible cross talk between agonist-induced association of translocated RhoA and translocated protein kinase C-alpha (PKC-alpha) and a role for heat shock protein 27 (HSP27) in mediating this interaction. Immunoprecipitation with HSP27 monoclonal antibody followed by immunoblotting with either RhoA antibody or PKC-alpha antibody indicated that acetylcholine induced associations of HSP27-RhoA and HSP27-PKC-alpha in the membrane fraction but not in the cytosolic fraction. Immunoprecipitation with anti-RhoA monoclonal antibody followed by immunoblotting with PKC-alpha antibody indicated that acetylcholine induced a significant complexing of RhoA-PKC-alpha in the membrane fraction but not in the cytosolic fraction. In summary, the data indicate that agonist-induced contraction is associated with 1) association of translocated RhoA with HSP27 on the membrane, 2) association of translocated PKC-alpha with HSP27 on the membrane, and 3) association of PKC-alpha with RhoA on the membrane. The data suggest an important role for HSP27 in modulating a multiprotein complex that includes translocated RhoA and PKC-alpha.
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Department of Pediatrics, University of Michigan Medical Center, Ann Arbor, Michigan 48109, USA. bitar@umich.edu
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D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015153:Blotting, Western; D002518:Ceramides / Q000494:pharmacology; D003106:Colon / Q000166:cytology / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000235:genetics / Q000502:physiology*; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000235:genetics / Q000502:physiology*; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000235:genetics / Q000502:physiology*; D051571:Protein Kinase C-alpha; D011817:Rabbits; D015398:Signal Transduction / Q000502:physiology; D014178:Translocation, Genetic / Q000235:genetics; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000502:physiology*
D000911:Antibodies, Monoclonal; D002518:Ceramides; D007527:Isoenzymes; D009363:Neoplasm Proteins; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D020742:rhoA GTP-Binding Protein; D000109:Acetylcholine