title: Phosphatidylinositol 3-kinase in bovine lens and its stimulation by insulin and IGF-1.
abstract: OBJECTIVE
  To identify and characterize phosphatidylinositol 3-kinase (PI-3K) in the lens and to study its involvement as a signal mediator in lens epithelial cells exposed to insulin and insulin-like growth factor (IGF)-1, which are known to induce lens epithelial cell proliferation and differentiation into fiber cells.
  
  
  METHODS
  Concentric fiber cell layers from single bovine lens were prepared by dissolution in buffer. PI-3K activity in capsule-epithelium and fiber cell layers was determined after immunoprecipitation with antibodies against p85, the regulatory subunit of PI-3K. High-performance liquid chromatography on an ion exchange column (Partisil-SAX; Whatman, Maidstone, United Kingdom) was used to identify PI-3K reaction products. Cultured bovine lens epithelial cells were stimulated with insulin or IGF-1, and PI-3K activity was determined after immunoprecipitation with antibody against phosphotyrosine. Association of p85 with other proteins after stimulation was determined in anti-p85 immunoprecipitates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western immunoblot analysis using anti-phosphotyrosine antibody.
  
  
  RESULTS
  PI-3K activity was found in both lens epithelial cells and fiber cells. The highest specific activity was found in the capsule-epithelium, but there was considerable activity in other fiber cell layers. Insulin and IGF-1 stimulated the PI-3K activity in epithelial cells in culture by more than 100%, and activation of the enzyme resulted in tyrosine phosphorylation of the p85 subunit. After stimulation, the p85 subunit of PI-3K was associated with 100- and 180-kDa tyrosine phosphorylated proteins.
  
  
  CONCLUSIONS
  The activation of PI-3K and its association with specific tyrosine-phosphorylated proteins may be important in insulin and IGF-1 signal transduction in lens epithelial cells. The presence of significant PI-3K activity throughout the lens further suggests that this signal transduction enzyme is sustained in fiber cells.
authors.affiliation: Louisiana State University Medical Center Eye Center and Neuroscience Center, New Orleans, Louisiana 70112, USA.
authors.forename: G
authors.initials: G
authors.lastname: Chandrasekher
authors.forename: H E
authors.initials: HE
authors.lastname: Bazan
mesh_terms: D000818:Animals; D002417:Cattle; D002454:Cell Differentiation; D002455:Cell Division; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects* / Q000201:enzymology; D007328:Insulin / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007908:Lens, Crystalline / Q000187:drug effects* / Q000201:enzymology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism
chemical_list: D007328:Insulin; D014443:Tyrosine; D007334:Insulin-Like Growth Factor I; D019869:Phosphatidylinositol 3-Kinases