TYROSINE PHOSPHORYLATION AND MAMMALIAN RETINA

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Signaling through intracellular tyrosine phosphorylation is an integral component of many normal and aberrant cellular processes including growth, differentiation, proliferation, maintenance and survival. The initial event in these signaling cascades in non-ocular tissues involves stimulus mediated activation of a tyrosine kinase. Our studies on tyrosine phosphorylation in mammalian retinas in vivo show that: 1) light stimulates tyrosine phosphorylation of photoreceptor out segment proteins, 2) rod outer segments (ROS) from light-adapted retinas are enriched with the tyrosine kinase c-Src,PLCgamma/1, and arresting, 3) at least three putative tyrosine kinase substrates are present in isolated ROS (Phospholipase C (PLCgamma/1), Phosphatidylinositol 3-kinase (PI 3-kinase) and the tyrosine phosphatase Syp) and 4) light induces a changes in the localization of PLCgamma/1 from the inner to the outer segment of photoreceptors that is coincident with increased tyrosine phosphorylation in ROS. Based on these observations, we hypothesize that light stimulates c-Src by inducing its autophosphorylation on tyrosine residues. The phosphotyrosine (P-Y) residues on C-Src can then associated with Src homology regions (SH2) domains present in other photoreceptor signaling proteins (e.g. PLCgamma/1, PI 3-kinase and the tyrosine phosphatase Syp) or with phosphotyrosine binding domains (PTB) on other proteins leading to tyrosine phosphorylation of these proteins and subsequent transport or translocation to the outer segment. The goals of the current application are to determine: 1. subsequent transport or translocation to the outer segment. The goals of the current application are to determine: 1. if c- Src is a light-activated tyrosine kinase in rat photoreceptor cells/ rod outer segment (ROS) in vivo; 2. the identities of the major protein substrates that are tyrosine phosphorylated by light in photoreceptor cells in vivo; and 3. if tyrosine phosphorylation in vivo promotes the translocation of specific proteins to photoreceptor ROS. Several different in vivo approaches including biochemical, immunological and cytochemical techniques will be utilized in addressing these specific aims. We propose that in vivo light-mediated tyrosine phosphorylation may play a role in the maintenance of the functional or structural polarization of photoreceptor cells which may contribute to the overall maintenance and survival of these cells.