PKC-gamma phosphorylation of connexin 46 in the lens cortex

To identify the role of PKC-gamma in control of and phosphorylation of connexin 46 (Cx46) in the lens cortex.

The association between PKC-gamma and Cx46 was determined by co-immunoprecipitation from whole lens. Phosphorylation of Cx46 and activity of PKC-gamma were determined using Western blots, PKC activity assays, and inhibition of PKC activity by addition of isoform-specific PKC pseudosubstrate inhibitors.

Co-localization of PKC-gamma and Cx46 was observed in the bow regions and cortical regions of rat lens. PKC-gamma was not observed in the nuclear region and Cx46 was not observed in the epithelial layer. PKC-alpha was not found in lens cortex or nuclear regions. PKC-gamma could be co-immunoprecipitated with Cx46 from lens cortical regions. Cx46 was phosphorylated on both serine and threonine. No tyrosine phosphorylation was observed. The PKC-gamma specific pseudosubstrate inhibitor caused a 73% inhibition of serine phosphorylation on Cx46 at 1 microM, and, 36% inhibition of threonine phosphorylation at the same concentration. Inhibition of phosphorylation of Cx46 with PKC-alpha pseudosubstrate inhibitor was not observed.

PKC-gamma may phosphorylate Cx46, primarily on serine in whole lens. A role for PKC-gamma in control of lens cortical gap junctions is suggested.