The Role of the Scaffolding Protein INAD in Localization of Signaling Complexes to te Rhabdomeres of Drosophil Photoreceptors

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Title: The Role of the Scaffolding Protein INAD in Localization of Signaling Complexes to te Rhabdomeres of Drosophil Photoreceptors
Author: Hahn, Adrienne R.
Abstract: Organization of proteins into macromolecular complexes is one way cells maximize the speed , specificity , and efficiency of signal transduction . In fruit fly photoreceptors InaD , a scaffolding protein containing 5 PDZ domains , organizes proteins involved in the visual signaling pathway into complexes within a microvillar stack of membranes known as the rhabdomere . Light activation of rhodopsin activates a signaling cascade via a Gq -coupled reaction that quickly opens Ca++ -selective TRP channels . Subsequent Ca++ influx activates eye protein kinase C (ePKC ) , and calmodulin , which in turn modulate the activity of other visual proteins . Mutants in which the InaD / TRP association has been disrupted (inaD215 ) phenocopy the delayed inactivation of mutants lacking ePKC , suggesting that one of the functions of InaD includes localizing ePKC to its downstream targets such as the TRP channel . There are currently two different models for scaffolding proteins : the "beads on a string" or "tethering" model where the order of the binding domains and their respective binding partners is unimportant , and the "specific quaternary structure" model where the specific stereochemical orientation of the domains is vital for proper signaling . The latter model also allows for allosteric regulation of binding . We assess the "beads on a string" vs . the "specific quaternary structure" model for InaD , a scaffolding protein found in the photoreceptor cells of fruit flies , by analyzing the characteristics of the light response in flies expressing two InaD constructs where the order of the PDZ domains has been shuffled . Based on biochemical and electrophysiological data on these mutants , we conclude that the "specific quaternary structure" model applies best to InaD . In addition , we investigate whether the inaD215 phenotype is due to displacement of ePKC from microdomains of calcium initiated by TRP channels , by calcium imaging of photoreceptors expressing visual proteins tagged with CaMgaroo , a Ca++ -sensitive derivative of yellow fluorescent protein . After conducting these experiments , we conclude that this is not the case . Rather , the inaD215 phenotype is most likely due to the inability of ePKC to phosphorylate TRP and and attenuate its activity . These results also support the "specific quaternary structure" model for InaD .
URI: http : / /hdl .handle .net /2152 .5 /267
Date: 2004-12-15

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The Role of the Scaffolding Protein INAD in Localization of Signaling Complexes to te Rhabdomeres of Drosophil Photoreceptors. Graduate School of Biomedical Sciences. Available electronically from http : / /hdl .handle .net /2152 .5 /267 .

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