In addition to their established role in the generation of second messengers by phospholipase C-catalysed hydrolysis, three of the inositol-containing phospholipids, and phosphatidylinositol, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-biphosphate are substrates for a second receptor-regulated enzyme, phosphoinositide 3-kinase. This ATP-dependent enzyme catalyses the phosphorylation of the D-3 OH group of each of the above lipids and is activated by a range of growth factors and hormones that stimulate receptor tyrosine kinases and guanine nucleotide-dependent regulatory protein [G-protein]-linked receptors in a diverse variety of target cells. Accumulation of the products of phosphoinositide 3-kinase, phosphatidylinositol 3-phosphate, phosphatidylinositol 3,4-bisphosphate and phosphatidylinositol 3,4,5- triphosphate is rapid in stimulated cells. It is probable that these 3- phosphorylated inositol lipids or some metabolite derived from them serves as an important signal in cellular responses to hormones and growth factors. Recent observations implicate phosphoinositide 3-kinase products as regulators of a calcium and phospholipid-dependent protein kinase, and modulators of protein secretion and trafficking. The long- term goal of the proposed research is to understand the cellular functions of receptor-stimulated formation of 3-phosphorylated inositol lipids. Two major questions will be addressed: of receptor-stimulated How do receptors, particularly those coupled to G-proteins, regulate phosphoinositide 3-kinase activity and what are the metabolic fates of the 3-phosphorylated inositol lipids when formed in stimulated cells. The specificity with which G-protein-coupled receptors stimulate the accumulation of 3-phosphorylated inositol lipids will be established. Cell lines expressing pathways for both tyrosine kinase receptor and G- protein-coupled receptor stimulation will be identified. The pathways by which these two classes of receptors regulate phosphoinositide 3- kinase will be compared in intact cells emphasizing the role of role of second messengers, protein kinases and G-proteins in this process. Using cell-free assays, the biochemical mechanisms by which G-protein-coupled receptors activate phosphoinositide 3-Kinase will be determined and the protein components of this system identified. The substrate-specificity of phosphoinositide 3-kinase and the metabolic interrelationships of the products of this enzyme and the other inositol-containing lipids will be examined in both intact and broken cell systems. The 3-phosphorylated inositol lipids will be prepared in radiolabeled and unlabeled forms. The metabolism of these lipids will be examined in cell and tissue homogenates. Successful completion of this program of research will greatly advance our understanding of the regulation and functions of phosphoinositide 3-kinase and its products, both of which promise to be pivotal components of an important signal transduction system that mediates cellular responses to stimulation by hormones and growth factors.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM050388-02
Application #
2188196
Study Section
Pharmacology A Study Section (PHRA)
Project Start
1994-08-01
Project End
1998-07-31
Budget Start
1995-08-01
Budget End
1996-07-31
Support Year
2
Fiscal Year
1995
Total Cost
Indirect Cost
Name
State University New York Stony Brook
Department
Pharmacology
Type
Schools of Medicine
DUNS #
804878247
City
Stony Brook
State
NY
Country
United States
Zip Code
11794
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Wu, Tao; Kooi, Craig Vander; Shah, Pritom et al. (2014) Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration. FASEB J 28:861-70
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Deevska, Gergana; Sunkara, Manjula; Karakashian, Claudia et al. (2014) Effect of procysteine on aging-associated changes in hepatic GSH and SMase: evidence for transcriptional regulation of smpd3. J Lipid Res 55:2041-52
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Subramanian, Thangaiah; Subramanian, Karunai Leela; Sunkara, Manjula et al. (2013) Syntheses of deuterium labeled prenyldiphosphate and prenylcysteine analogues for in vivo mass spectrometric quantification. J Labelled Comp Radiopharm 56:370-5
Karapetyan, Anush V; Klyachkin, Yuri M; Selim, Samy et al. (2013) Bioactive lipids and cationic antimicrobial peptides as new potential regulators for trafficking of bone marrow-derived stem cells in patients with acute myocardial infarction. Stem Cells Dev 22:1645-56
Mitra, Mayurranjan S; Chen, Zhouji; Ren, Hongmei et al. (2013) Mice with an adipocyte-specific lipin 1 separation-of-function allele reveal unexpected roles for phosphatidic acid in metabolic regulation. Proc Natl Acad Sci U S A 110:642-7
Onono, Fredrick; Subramanian, Thangaiah; Sunkara, Manjula et al. (2013) Efficient use of exogenous isoprenols for protein isoprenylation by MDA-MB-231 cells is regulated independently of the mevalonate pathway. J Biol Chem 288:27444-55

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