Association of INAD with NORPA is essential for controlled activation and deactivation of Drosophila phototransduction in vivo

• DOI: 10.1073/pnas.94.23.12682
• Publication Date: 2002-07-26T14:31:44Z
• Authors: Bih-Hwa Shieh; Mei-Ying Zhu; John K. Lee; Ingrid M. Kelly; Frohar Bahiraei
• Keywords: Animals, Genetically Modified; 0301 basic medicine; 03 medical and health sciences; Phosphatidylinositol Diacylglycerol-Lyase; Type C Phospholipases; Phospholipase C beta; Animals; Drosophila Proteins; Drosophila; Eye Proteins; Vision, Ocular; Signal Transduction

Visual transduction in Drosophila is a G protein-coupled phospholipase C-mediated process that leads to depolarization via activation of the transient receptor potential (TRP) calcium channel. Inactivation-no-afterpotential D (INAD) is an adaptor protein containing PDZ domains known to interact with TRP. Immunoprecipitation studies indicate that INAD also binds to eye-specific protein kinase C and the phospholipase C, no-receptor-potential A (NORPA). By overlay assay and site-directed mutagenesis we have defined the essential elements of the NORPA–INAD association and identified three critical residues in the C-terminal tail of NORPA that are required for the interaction. These residues, Phe-Cys-Ala, constitute a novel binding motif distinct from the sequences recognized by the PDZ domain in INAD. To evaluate the functional significance of the INAD–NORPA association in vivo , we generated transgenic flies expressing a modified NORPA, NORPA

C1094S

, that lacks the INAD interaction. The transgenic animals display a unique electroretinogram phenotype characterized by slow activation and prolonged deactivation. Double mutant analysis suggests a possible inaccessibility of eye-specific protein kinase C to NORPA

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, undermining the observed defective deactivation, and that delayed activation may similarly result from NORPA

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being unable to localize in close proximity to the TRP channel. We conclude that INAD acts as a scaffold protein that facilitates NORPA–TRP interactions required for gating of the TRP channel in photoreceptor cells.