User talk:Cojenho

PROPOFOL ACTIONS ON PROTEINS KINASE Ca TRANSLOCATION AND ACTIVATION

ABSTRACT

Protein kinase C (PKC), a key element in control of cell signaling has been suggested to be one of the potential targets for general anesthetics. In this study, we investigate the effect of propofol (2.6-diisopropyl phenol), a common intravenous anesthetic, on PKCa membrane translocation and potential direct interaction with PKC in vitro and in cellular milieu. It was found that propofol significantly enhanced PKCa membrane translocation and activation in the presence of PKC cofactors, TPA/DAG, PS and Ca2+. In the absence of PKC activator, higher concentration of propofol was sufficient to translocate PKCa, but these PKCa were enzymatic inactive. Strong evidence of propofol modulating PKC-phorbol ester interaction through PKC C1 domain was obtained using fluorescent phorbol ester, SAPD, suggesting propofol directly interacts with PKCa at C1 domain and potentiation of membrane translocation is not mediated through membrane lipids, while membrane lipids might stabilize this interaction. Fluorescence imaging of GFP-tagged PKCa transfected in Chinese Hamster Ovarian epithelial cells (CHO-K1) cells demonstrated propofol potentiated PKCa membrane translocation and, in the absence of activator, propofol progressively translocated PKCa to plasma membrane as well as intracellular structures in a Ca2+ dependent manner, in consistent with in vitro results from fluorescence anisotropy measurements. GFP-PKC mutants with C1a deletion transfected in CHO-K1 cells revealed a non-essential role of C1a for propofol effect. The findings suggest propofol effect on PKCa cellular movement and functional consequences is likely via C1b domain interactions, and strengthen the proposition of PKC involvement in the actions of propofol and possibly other general anesthetics.