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A.S. Menko, L. Zhang; Src Kinase Regulation of 6 Integrin Signaling in Lens Cell Differentiation . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1255.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We have previously shown that α6 integrin plays a requisite role in lens cell differentiation and that this integrin recruits its downstream signaling effectors in a differentiation specific manner. In this study we investigate whether Src family kinases regulate this α6 integrin signaling pathway and examine their role lens cell differentiation. Methods: Primary quail lens epithelial cells were grown in culture until they formed differentiated lentoid structures. The composition of α6 integrin signaling complexes was determined by immunoprecipitation of cell extracts with antibody to α6 integrin followed by immunoblotting with antibodies to candidate signaling molecules. These included the growth factor receptor IGF1-R, the adaptor protein Shc, and its downstream signaling effector the MAP kinase ERK. The activation state of signaling molecules in α6 integrin complexes was determined by immunoblot analysis with phosphospecific antibodies. To assess whether recruitment of signaling proteins to α6 integrin complexes and their activation was dependent on Src family kinase activity, lens cultures were grown in the presence of the Src-specific inhibitor PP1. Results: Our previous studies show that culturing lens epithelial cells in the presence of the Src kinase inhibitor PP1 promotes cell cycle withdrawal and initiation of lens cell differentiation. We now report that continued maintenance of lens cultures in the Src inhibitor prevented the formation of lentoids. This result demonstrated that Src kinases play distinct roles at different stages in the process of lens cell differentiation. We next investigated whether the mechanism by which PP1 inhibited lentoid formation involved its blocking of the α6 integrin signaling pathway. In normal differentiating lens cell cultures components of the α6 integrin signaling complex included activated IGF-1R, phosphoShc, and activated ERK. Exposure of lens cultures to the PP1 Src inhibitor resulted in diminished phosphorylation of IGF-1R, Shc and ERK in the α6 integrin protein complexes. These results demonstrate that Src kinase activity is required for α6 integrin signaling in differentiating lens cells. Conclusions: Src kinase activity is required for lentoid formation as well as for α6 integrin activation of IGF-1R and their downstream signaling effectors Shc and ERK. These results show that Src kinases play an important role in lens cell differentiation as mediators of α6 integrin signaling.
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