Growth Factor Receptor-Bound Protein 14 Undergoes Light-Dependent Intracellular Translocation in Rod Photoreceptors: Functional Role in Retinal Insulin Receptor Activation

Abstract

Growth factor receptor-bound protein 14 (Grb14) is involved in growth factor receptor tyrosine kinase signaling. Here we report that light causes a major redistribution of Grb14 among the individual subcellular compartments of the retinal rod photoreceptor. Grb14 is localized predominantly to the inner segment, nuclear layer and synapse in dark-adapted rods, whereas in the light-adapted rods, Grb14 redistributed throughout the entire cell, including the outer segment. The translocation of Grb14 requires photoactivation of rhodopsin, but not signaling through the phototransduction cascade, and is not based on direct Grb14-rhodopsin interactions. We previously hypothesized that Grb14 protects light-dependent insulin receptor (IR) activation in rod photoreceptors against dephosphorylation by protein tyrosine phosphatase 1B. Consistent with this hypothesis, we failed to observe light-dependent IR activation in Grb14 −/− mouse retinas. Our studies suggest that Grb14 translocates to photoreceptor outer segments after photobleaching of rhodopsin and protects IR phosphorylation in rod photoreceptor cells. These results demonstrate that Grb14 can undergo subcellular redistribution upon illumination and suggest that rhodopsin photoexcitation may trigger signaling events alternative to the classical transducin activation.Phosphoinositide 3-kinase kinase (PI3K) is at the heart of one of the major signal transduction pathways (1-4). The signals mediated by this enzyme influence a wide variety of cellular functions, including cell growth, differentiation and survival, glucose metabolism and cytoskeletal organization. The PI3K is expressed in photoreceptor cells and is regulated through the light-induced tyrosine phosphorylation of the insulin receptor (IR) in vivo (5;6). We have reported that light-induced tyrosine phosphorylation of IR requires the photoactivation of rhodopsin, but not transducin signaling (7). We also found that Address Correspondence to: Raju V.S. Rajala, University of Oklahoma Health Sciences Center, 608 Stanton L. Young Blvd., Oklahoma City, OK 73104. photoreceptor-specific deletion of IR resulted in stress-induced photoreceptor degeneration, suggesting the importance of IR in the survival of photoreceptor neurons (8).The molecular mechanism behind the light-induced activation of retinal IR is not known. Retinal IR has a high basal level of autophosphorylation compared to liver IR (9) and retinal IR autophosphorylation is light-dependent (5). These observations led us to hypothesize that retinal IR phosphorylation could be modulated by soluble factor(s) in the retina. To identify the regulators of IR, yeast two-hybrid screening of a bovine retinal cDNA library with the cytoplasmic domain of retinal IR (10) identified growth factor receptor-bound protein 14 (Grb14) (11;12), which binds to various tyrosine kinase receptors including IR (13-16). The crystal structure of the tyrosine kinase domain in complex with the IR-interacting domain of Grb14 has been resolved and revealed that Grb14 acts as a ...

Phosphoinositide 3-kinase signaling in the vertebrate retina

“…Previously, we hypothesized that Grb14 protects lightdependent IR activation in rod photoreceptors against dephosphorylation by PTP1B ( 49 ). We failed to observe light-dependent IR activation in Grb14 knockout mouse retinas, supporting our hypothesis ( 162 ). Our studies suggest that Grb14 translocates to ROSs after photobleaching of rhodopsin and protects IR phosphorylation in rod photoreceptor cells.…”

“…To identify the regulators of IR, yeast two-hybrid screening of a bovine retinal cDNA library constructed with the cytoplasmic domain of retinal IR ( 68 ) identifi ed growth factor receptor-bound protein 14 (Grb14) ( 156,157 ), which binds to various tyrosine kinase receptors including IR (158)(159)(160)(161). Grb14 is localized predominantly in the rod inner segment, nuclear layer, and synapse in dark-adapted rods, whereas Grb14 is redistributed throughout the entire cell, including the outer segment, in the light-adapted rods ( 162 ). The translocation of Grb14 requires photoactivation of rhodopsin, but PI3K SIGNALING IN DIABETIC RETINOPATHY PI3K and Akt are downstream effectors of insulin signaling ( 136 ) as well as important signaling molecules in the regulation of glycogen metabolism in myocytes, lipocytes, and hepatocytes ( 137,138 ).…”

“…PI3K/Akt can regulate the early steps of de novo synthesis by modulating phosphoribosylpyrophosphate production by the nonoxidative pentose phosphate pathway and the late steps by modulating activity of the bifunctional enzyme aminoimidazole-carboxamide ribonucleotide transformylase IMP cyclohydrolase ( 178 ). These studies defi ned not signaling through the phototransduction cascade, and is not based on direct Grb14-rhodopsin interactions ( 162 ). Previously, we hypothesized that Grb14 protects lightdependent IR activation in rod photoreceptors against dephosphorylation by PTP1B ( 49 ).…”

Phosphorylated Grb14 Is an Endogenous Inhibitor of Retinal Protein Tyrosine Phosphatase 1B, and Light-Dependent Activation of Src Phosphorylates Grb14

“…A high expression of Grb14 in myocardial tissue activates the phosphoinositide 3-kinase (PI3K)-Akt pathway, and ablation of Grb14 resulted in myocardial infarction and decreased PI3K/Akt activation (28). In the retina, light induces activation of the IR, and ablation of Grb14 results in the loss of light-dependent IR activation (31). The IR activation is essential for photoreceptor neuron survival (29,32).…”

“…NJ). Generation of a polyclonal Grb14 antibody was described earlier (31). All other reagents were of analytical grade from Sigma (St. Louis, MO).…”