Nutritional vitamin A depletion has been known for a century to cause a complex deficiency syndrome that affects multiple organs. Lack of retinoic acid (RA) leading to faulty transcription is the best studied mechanism. However, deficiencies in development, growth, reproduction and immunity were consequences of absence of vitamin A itself, since RA could not fully reverse these symptoms. We and others have described retinoid pathways that operate in the cytoplasm, independently of transcription. Further, we have identified serine/threonine kinases of the cRaf and PKC families as direct targets of vitamin A action. Vitamin A binds their regulatory domains and bound vitamin A sensitizes these kinases for redox-mediated activation. Redox activation refers to the alternative pathway operating through reactive oxygen species {ROS) as second messenger. The main objective is to prove the essential role that vitamin A plays in redox signaling, by establishing cause /effect relationships. Because ROS activates several kinases at once we propose to study the vitamin A dependence of two isoforms, alpha and theta, separately using a genetic approach. We will map the vitamin A binding site by scanning mutagenesis, already successful with cRaf (AIM #1). Using mutant PKCalpha and theta where retinolbinding sites are deleted, the biological function will be probed in vivo and in vitro with transgenic, as well as knock-in, cell lines and mice. IL-2 production and T cell proliferation are PKCtheta-dependent immunologically significant processes (AIM # 2). We hypothesize that ROS with help of vitamin A produces covalent modifications, leading to disassembly of the zinc-finger as prelude to kinase unfolding and activation.
AIM # 3 is devoted to biochemical analyses of pertinent cysteine-modifications.
Aim # 4 will focus on retinol as co-factor in mediating the release of zinc and conformation change in the zinc-finger consequent to redox activation, using intra-vital imaging. The results will establish a new mechanism of action of vitamin A, help understand vitamin A deficiency syndromes in reproduction, development and immunity, and open new approaches to prevention and treatment of cancer.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK069348-05
Application #
7633156
Study Section
Integrative Nutrition and Metabolic Processes Study Section (INMP)
Program Officer
Maruvada, Padma
Project Start
2005-09-15
Project End
2011-06-30
Budget Start
2009-07-01
Budget End
2011-06-30
Support Year
5
Fiscal Year
2009
Total Cost
$339,349
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
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Acin-Perez, Rebeca; Hoyos, Beatrice; Gong, Jianli et al. (2010) Regulation of intermediary metabolism by the PKCdelta signalosome in mitochondria. FASEB J 24:5033-42
Chiu, Haw-Jyh; Fischman, Donald A; Hammerling, Ulrich (2008) Vitamin A depletion causes oxidative stress, mitochondrial dysfunction, and PARP-1-dependent energy deprivation. FASEB J 22:3878-87
Rogge, Barbara; Itagaki, Yasuhiro; Fishkin, Nathan et al. (2005) Retinoylserine and retinoylalanine, natural products of the moth Trichoplusia ni. J Nat Prod 68:1536-40