The objective of our research is to gain insight into how protein kinase C (PKC) regulates G-protein coupled phospholipase CB-mediated signal transduction. This signaling mechanism is widely employed in the central nervous system for the action of several neurotransmitters including glutamate, serotonin, acetylcholine, norepinephrine, and histamine. Activation of phospholipase CB-3 leads to generation of two messengers: diacylglycerol (DAG) and inositol trisphosphate (1P3). IP3 mobilizes intracellular calcium whereas DAG activates PKC. PKC has been implicated in regulation of signal transduction as well as diverse cellular processes including growth, differentiation, and tumorigenesis. We will use the visual transduction in Drosophila melanogaster as a model system. In the visual cascade, E photoreceptor-specific protein kinase C (eye-PKC) is involved in desensitization, deactivation and light adaptation of the visual signaling. We previously showed that eye-PKC is tethered to a signal transduction complex by interacting with an adaptor protein INAD. A lack of the INAD-eyePKC interaction leads to a loss of in vivo activity of eye-PKC. Moreover, we demonstrated that INAD and TRP, which also associate with INAD, are phosphorylated by eye-PKC. We will: (1) identify eye-PKC phosphorylation sites in INAD; (2) investigate how phosphorylation of INAD affects protein-protein interactions with other components of the signaling complex; (3) identify eye-PKC phosphorylation sites in TAP; (4) investigate how phosphorylation of TRP may modulate the channel activities and protein-protein interaction with INAD; and (5) investigate if NORPA is also a substrate of eye-PKC and how NORPA is regulated by phosphorylation. PKC regulates many facets of cellular function. We believe that our research on regulation of G-protein coupled signaling by PKC will provide insight into modulation of cell-cell communications in the nervous system. Un-regulated signaling may lead to hyperactivity or degeneration of neurons, which has been implicated in mans pathophysiological conditions such as epilepsy and neurodegenerative diseases.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY009743-12
Application #
6874842
Study Section
Molecular, Cellular and Developmental Neurosciences 2 (MDCN)
Program Officer
Mariani, Andrew P
Project Start
1993-04-01
Project End
2007-02-28
Budget Start
2005-04-01
Budget End
2006-03-31
Support Year
12
Fiscal Year
2005
Total Cost
$339,750
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Pharmacology
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Lu, Haiqin; Leung, Hung-Tat; Wang, Ning et al. (2009) Role of Ca2+/calmodulin-dependent protein kinase II in Drosophila photoreceptors. J Biol Chem 284:11100-9
Wang, Ning; Leung, Hung-Tat; Pak, William L et al. (2008) Role of protein phosphatase 2A in regulating the visual signaling in Drosophila. J Neurosci 28:1444-51
Peng, Li; Popescu, Daniela C; Wang, Ning et al. (2008) Anchoring TRP to the INAD macromolecular complex requires the last 14 residues in its carboxyl terminus. J Neurochem 104:1526-35
Popescu, Daniela C; Ham, Amy-Joan L; Shieh, Bih-Hwa (2006) Scaffolding protein INAD regulates deactivation of vision by promoting phosphorylation of transient receptor potential by eye protein kinase C in Drosophila. J Neurosci 26:8570-7
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Shieh, Bih-Hwa; Parker, Lisan; Popescu, Daniela (2002) Protein kinase C (PKC) isoforms in Drosophila. J Biochem 132:523-7
Kumar, R; Shieh, B H (2001) The second PDZ domain of INAD is a type I domain involved in binding to eye protein kinase C. Mutational analysis and naturally occurring variants. J Biol Chem 276:24971-7
Liu, M; Parker, L L; Wadzinski, B E et al. (2000) Reversible phosphorylation of the signal transduction complex in Drosophila photoreceptors. J Biol Chem 275:12194-9
Adamski, F M; Zhu, M Y; Bahiraei, F et al. (1998) Interaction of eye protein kinase C and INAD in Drosophila. Localization of binding domains and electrophysiological characterization of a loss of association in transgenic flies. J Biol Chem 273:17713-9
Shieh, B H; Zhu, M Y; Lee, J K et al. (1997) Association of INAD with NORPA is essential for controlled activation and deactivation of Drosophila phototransduction in vivo. Proc Natl Acad Sci U S A 94:12682-7

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