PKC-gamma phosphorylation of connexin 46 in the lens cortex.
PKC-gamma phosphorylation of connexin 46 in the lens cortex.
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DOI：
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发表时间：
2001-10
期刊：
影响因子：
2.2
通讯作者：
S. Saleh;L. Takemoto;D. Zoukhri;D. Takemoto
中科院分区：
文献类型：
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作者：
S. Saleh;L. Takemoto;D. Zoukhri;D. Takemoto
目的研究蛋白激酶C-γ（PKC-γ）在透镜皮质连接蛋白46（Cx46）磷酸化和调控中的作用。方法采用免疫共沉淀法从全透镜中检测PKC-γ与Cx46的结合。使用蛋白质印迹、PKC活性测定和通过加入亚型特异性PKC假底物抑制剂抑制PKC活性来测定Cx46的磷酸化和PKC-γ的活性。结果在大鼠透镜的弓状区和皮质区存在PKC-γ和Cx46的共定位。PKC-γ在细胞核区未观察到，Cx46在上皮层未观察到。在透镜皮质和核区未发现PKC-α。PKC-γ可与透镜皮质区的Cx46共沉淀。Cx46在丝氨酸和苏氨酸上都被磷酸化。未观察到酪氨酸磷酸化。PKC-γ特异性假底物抑制剂在1 μ M时对Cx46上丝氨酸磷酸化的抑制率为73%，在相同浓度下对苏氨酸磷酸化的抑制率为36%。未观察到PKC-α假底物抑制剂对Cx46磷酸化的抑制。结论PKC-γ可使整个透镜中的Cx46磷酸化，主要作用于丝氨酸。PKC-γ在控制透镜皮质间隙连接中的作用。
PURPOSE To identify the role of PKC-gamma in control of and phosphorylation of connexin 46 (Cx46) in the lens cortex. METHODS 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. RESULTS 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. CONCLUSIONS 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.