In vivo regulation of phosphoinositide 3-kinase in retina through light-induced tyrosine phosphorylation of the insulin receptor beta-subunit.
Recently, we have shown that phosphoinositide 3-kinase (PI3K) in bovine rod outer segment (ROS) is activated in vitro by tyrosine phosphorylation of the C-terminal tail of the insulin receptor (Rajala, R. V. S., and Anderson, R. E. (2001) Invest. Ophthal. Vis. Sci. 42, 3110-3117). In this study, we have investigated the in vivo mechanism of PI3K activation in the rodent retina and report the novel finding that light stimulates tyrosine phosphorylation of the beta-subunit of the insulin receptor (IRbeta) in ROS membranes, which leads to the association of PI3K enzyme activity with IRbeta. Retinas from light- or dark-adapted mice and rats were homogenized and immunoprecipitated with antibodies against phosphotyrosine, IRbeta, or the p85 regulatory subunit of PI3K, and PI3K activity was measured using PI-4,5-P(2) as substrate. We observed a light-dependent increase in tyrosine phosphorylation of IRbeta and an increase in PI3K enzyme activity in isolated ROS and in anti-phosphotyrosine and anti-IRbeta immunoprecipitates of retinal homogenates. The light effect was localized to photoreceptor neurons and is independent of insulin secretion. Our results suggest that light induces tyrosine phosphorylation of IRbeta in outer segment membranes, which leads to the binding of p85 through its N-terminal Src homology 2 domain and the generation of PI-3,4,5-P(3). We suggest that the physiological role of this process may be to provide neuroprotection of the retina against light damage by activating proteins that protect against stress-induced apoptosis.
Rajala
Raju V S
RV
Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA. raju-rajala@ouhsc.edu
McClellan
Mark E
ME
Ash
John D
JD
Anderson
Robert E
RE
D000818:Animals; D002417:Cattle; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D010857:Pigment Epithelium of Eye / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011972:Receptor, Insulin / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012160:Retina / Q000201:enzymology / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000201:enzymology / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D019000:Phosphotyrosine; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D011972:Receptor, Insulin