Phosphoinositides: Minor Lipids Make a Major Impact on Photoreceptor Cell Functions

Activation of the phosphoinositide (PI) cycle generates the second messengers that control various aspects of cellular signaling. We have previously shown that two PI cycle enzymes, type II phosphatidylinositol 5-phosphate 4-kinase (PIPK IIα) and phosphoinositide 3-kinase (PI3K), are activated through light stimulation. In our earlier studies, we measured enzyme activities, instead of directly measuring the products, due to lack of sensitive analytical techniques. Cells have very low levels of PIs, compared to other lipids, so special techniques and sensitive analytical instruments are necessary for their identification and quantification. There are also other considerations, such as different responses in different cell types, which may complicate quantification of PIs. For example, although light activated PIPK IIα, there was no increase in PI-4,5-P2 measured by liquid chromatography-mass spectrometry (LC/MS) This discrepancy is due to the heterogeneous nature of the retina, which is composed of various cell types. In this study, we examined PI generation in situ using immunohistochemistry with specific PI antibodies. PIs were generated in specific retinal cell layers, suggesting that analyzing PIs from the total retina by LC/MS underscores the significance. This suggests that PI-specific antibodies are useful tools to study the cell-specific regulation of PIs in the retina

Insulin receptor regulates photoreceptor CNG channel activity

Photoreceptor cyclic nucleotide gated (CNG) channels are critical elements in phototransduction and light adaptation. Here we report that insulin receptor (IR), an integral membrane protein, directly phosphorylates the CNGA1 subunit of CNG channels that in turn affects the function of these channels negatively. The IR phosphorylates Tyr⁴⁹⁸ and Tyr⁵⁰³ residues on CNGA1 that are situated at the membrane-cytoplasmic interface. The IR tyrosine kinase activity is essential for the inhibition of CNG channel. To maintain the channels in an off state, it is necessary not only to have a precise balance of the cGMP levels but also to have a control on the cGMP sensitivity of the CNG channels itself. In this study, we observed that the channel opens at a lower concentration of cGMP in IR⁻/⁻ mice. These studies suggest that IR regulates the modulation of CNG channel activity in vivo.10 page(s

Light activation of the insulin receptor regulates mitochondrial hexokinase. A possible mechanism of retinal neuroprotection

The serine/threonine kinase Akt has been shown to mediate the anti-apoptotic activity through hexokinase(HK)–mitochondria interaction. We previously reported that Akt activation in retinal rod photoreceptor cells is mediated through the light-dependent insulin receptor (IR)/PI3K pathway. Our data indicate that lightinduced activation of IR/PI3K/Akt results in the translocation of HK-II to mitochondria. We also found that PHLPPL, a serine/threonine phosphatase, enhanced the binding of HK-II to mitochondria. We found a mitochondrial targeting signal in PHLPPL and our study suggests that Akt translocation to mitochondria could be mediated through PHLPPL. Our results suggest that the light-dependent IR/PI3K/Akt pathway regulates hexokinase– mitochondria interaction in photoreceptors. Down-regulation of IR signaling has been associated with ocular diseases of retinitis pigmentosa, diabetic retinopathy, and Leber Congenital Amaurosis-type 2, and agents that enhance the binding interaction between hexokinase and mitochondria may have therapeutic potential against these ocular diseases.11 page(s

Growth factor receptor-bound protein 14 : a new modulator of photoreceptor-specific cyclic-nucleotide-gated channel

Growth factor receptor-bound protein 14 (Grb14) is an adaptor protein that is involved in receptor tyrosine kinase signalling. In this study, we report that Grb14 interacts with the rod photoreceptor-specific cyclic-nucleotide-gated channel alpha subunit (CNGA1) and decreases its affinity for cyclic guanosine monophosphate. Channel modulation is controlled by direct binding of the Grb14 Ras-associating domain with the carboxy-terminal region of CNGA1. We observed that the channel remains open in Grb14-mice that are exposed to light, suggesting that Grb14 is a normal physiological modulator of CNG channel function in vivo.7 page(s

Insulin receptor signaling regulates actin cytoskeletal organization in developing photoreceptors

The insulin receptor (ir) and ir signaling proteins are widely distributed throughout the cns. ir signaling provides a trophic signal for transformed retinal neurons in culture and we recently reported that deletion of ir in rod photoreceptors by cre/lox system resulted in stress‐induced photoreceptor degeneration. these studies suggest a neuroprotective role of ir in rod photoreceptor cell function. however, there are no studies available on the role of insulin‐induced ir signaling in the development of normal photoreceptors. to examine the role of insulin‐induced ir signaling, we analyzed cultured neuronal cells isolated from newborn rodent retinas. in insulin‐lacking cultures, photoreceptors from wild‐type rat retinas exhibited an abnormal morphology with a wide axon cone and disorganization of the actin and tubulin cytoskeleton. photoreceptors from ir knockout mouse retinas also exhibited a similar abnormal morphology. a novel finding in this study was that addition of docosahexaenoic acid, a photoreceptor trophic factor, restored normal axonal outgrowth in insulin‐lacking cultures. these data suggest that ir signaling pathways regulate actin and tubulin cytoskeletal organization in photoreceptors; they also imply that insulin and docosahexaenoic acid activate at least partially overlapping signaling pathways that are essential for the development of normal photoreceptors.Fil: Rajala, Raju V. S.. University of Oklahoma Health Sciences Center; Estados UnidosFil: Rajala, Ammaji. University of Oklahoma Health Sciences Center; Estados UnidosFil: Brush, Richard S.. University of Oklahoma Health Sciences Center; Estados UnidosFil: Rotstein, Nora Patricia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; ArgentinaFil: Politi, Luis Enrique. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentin