Metabolic features of photoreceptors, Mller cells and retinal pigment epithelium cells are strikingly different. Metabolic relationships between these cells are important for retina function and survival. A key component of this metabolic ecosystem is the extraordinary efficiency with which photoreceptors in the outer retina convert glucose to lactate. Highly efficient glycolysis of glucose to lactate in the presence of abundant O2 and abundant mitochondria is referred to as ?Aerobic glycolysis? or the ?Warburg effect?. It is a metabolic feature of retinas and also of many types of cancer cells. The molecular mechanism responsible for the Warburg effect is unknown. We will use strategies described in the following three specific aims to identify the molecular mechanisms that enhance aerobic glycolysis in retinas. In the first specific aim of this proposal we will measure metabolic flux in live cells to reveal which steps in glycolysis are enhanced in retinas. Our preliminary findings show that the steady state concentrations of glycolytic intermediates in retinas is very low and that flux through those intermediates is very fast. Our preliminary findings suggest that steps that involve production and consumption of NADH appear to be enhanced. In the second aim we will compare retinal and RPE proteins using mass spectrometry without and with cross-linking to determine how retinas favor reduction over oxidation of pyruvate. These experiments will demonstrate the relative enrichment of glycolytic vs. mitochondrial proteins in the retina and RPE. The cross-linking studies will identify protein-protein interactions between the enzymes that catalyze reactions in the glycolytic pathway. In the third aim we will use in vitro assays of glycolytic enzyme activity to determine how retinas favor reduction over oxidation of pyruvate in mitochondria. Assaying production of lactate from glucose in the presence of enzymes that can compete for consumption of glycolytic intermediates will test the hypothesis that glycolysis is enhanced by channeling of glycolytic products and substrates between enzymes.

Public Health Relevance

Our recent findings show that the steady state concentrations of glycolytic intermediates is low in the retina while metabolic flux through glycolysis is very fast. A previous study showed that enhancing glycolysis in photoreceptors can make them more robust to stress. Our findings will identify the mechanism that enhances glycolysis in photoreceptors and may reveal new strategies for slowing or preventing retinal degenerations. !

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
2R01EY006641-34
Application #
9814976
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Neuhold, Lisa
Project Start
1986-07-01
Project End
2023-11-30
Budget Start
2019-12-15
Budget End
2020-11-30
Support Year
34
Fiscal Year
2020
Total Cost
Indirect Cost
Name
University of Washington
Department
Biochemistry
Type
Schools of Medicine
DUNS #
605799469
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
Hurley, James B (2017) Warburg's vision. Elife 6:
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:
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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