Vertebrate photoreceptors are excited by illumination, from which they recover when illumination is reduced and adapt to steady light. Biochemical and electrophysiological studies have revealed an enzymatic cascade involving cyclic GMP that mediates phototransduction. Each step of the cascade is regulated by molecular mechanisms that allow recovery and adaptation. Ca2+ also plays an important role in phototransduction. For example, light-stimulated hydrolysis of cyclic GMP lowers intracellular [Ca2+] which then stimulates phosphorylation of rhodopsin and activation of guanylyl cyclase. The applicant proposes the following three specific aims in order to further understand the molecular aspects of such photoreceptor transduction cascades.
Specific Aim I is to investigate the mechanism by which photoactivated rhodopsin stimulates rhodopsin kinase to phosphorylate other rhodopsins, a process referred to as high gain phosphorylation. The effect of recoverin on high-gain phosphorylation will also be investigated.
Specific Aim II is to identify functional domains of photoreceptor membrane guanylyl cyclases. These studies focus on identifying interactions between the cyclase and a Ca2+-sensitive activator protein. The role of the kinase homology domain within the cyclase and the role of phosphorylation of the cyclase will also be investigated.
Specific Aim III is to identify proteins that are specifically expressed in a particular type of photoreceptor. Differential display polymerase chain reaction will be used with individual rod and cone cells to identify mRNA transcripts that are expressed in one photoreceptor type but not in another. The rationale for this approach is that proteins important to photoreceptor development and function will be identified.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY006641-14
Application #
2888234
Study Section
Special Emphasis Panel (ZRG1-VISB (01))
Project Start
1986-07-01
Project End
2001-06-30
Budget Start
1999-07-01
Budget End
2000-06-30
Support Year
14
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Washington
Department
Biochemistry
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Du, Jianhai; An, Jie; Linton, Jonathan D et al. (2018) How Excessive cGMP Impacts Metabolic Proteins in Retinas at the Onset of Degeneration. Adv Exp Med Biol 1074:289-295
Rajala, Ammaji; Wang, Yuhong; Brush, Richard S et al. (2018) Pyruvate kinase M2 regulates photoreceptor structure, function, and viability. Cell Death Dis 9:240
Zhu, Siyan; Yam, Michelle; Wang, Yekai et al. (2018) Impact of euthanasia, dissection and postmortem delay on metabolic profile in mouse retina and RPE/choroid. Exp Eye Res 174:113-120
Chao, Jennifer R; Knight, Kaitlen; Engel, Abbi L et al. (2017) Human retinal pigment epithelial cells prefer proline as a nutrient and transport metabolic intermediates to the retinal side. J Biol Chem 292:12895-12905
Kanow, Mark A; Giarmarco, Michelle M; Jankowski, Connor Sr et al. (2017) Biochemical adaptations of the retina and retinal pigment epithelium support a metabolic ecosystem in the vertebrate eye. Elife 6:
Hurley, James B (2017) Warburg's vision. Elife 6:
Du, Jianhai; Rountree, Austin; Cleghorn, Whitney M et al. (2016) Phototransduction Influences Metabolic Flux and Nucleotide Metabolism in Mouse Retina. J Biol Chem 291:4698-710
Du, Jianhai; Yanagida, Aya; Knight, Kaitlen et al. (2016) Reductive carboxylation is a major metabolic pathway in the retinal pigment epithelium. Proc Natl Acad Sci U S A 113:14710-14715
Zhang, Lijuan; Du, Jianhai; Justus, Sally et al. (2016) Reprogramming metabolism by targeting sirtuin 6 attenuates retinal degeneration. J Clin Invest 126:4659-4673
Contreras, Laura; Ramirez, Laura; Du, Jianhai et al. (2016) Deficient glucose and glutamine metabolism in Aralar/AGC1/Slc25a12 knockout mice contributes to altered visual function. Mol Vis 22:1198-1212

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