The basic goal of this study is to make a significant contribution to the understanding of the cellular mechanism of insulin action and the regulation of cellular metabolism in general. A comprehension of the mechanism of insulin action is crucial both from a basic science point of view and from a practical standpoint to understand the pathophysiology and to potentially design new methods of treatment of diabetes mellitus. The glucose transport protein will be the focus of the investigation. Specifically, the role of Ca2+- and phospholipid-sensitive protein kinase (protein kinase C, also known as Ca, PL-PK) in insulin mediated phosphorylation of the glucose transporter will be investigated. Recent evidence indicates that the glucose transport protein is phosphorylated by both insulin and protein kinase C. It is my hypothesis that phosphorylation of the glucose transport protein regulates glucose transport and the mechanism of insulin-induced phosphorylation of the glucose transporter may be mediated via protein kinase C. The experimental approach includes the following specific aims: i) characterization of phosphorylation of the glucose transport protein in isolated adipocyte plasma membranes and microsomes, ii) characterization of changes in phosphorylation of the glucose transporter in plasma membranes and microsomes from adipocytes pre-treated with insulin or activators (e.g., phospholipase C, diacylglycerol, the phorbol ester tetradecanoylphorbol acetate) and inhibitors (e.g., gossypol, quercitin, polymyxin B) of protein kinase C and iii) investigation of the effects of the above agents on phosphorylation of the glucose transport protein in intact adipocytes. In all experiments, changes in phosphorylation of the glucose transport protein will be correlated with changes in glucose transport activity and 3H cytochalasin B binding to the glucose transporter. These studies should add considerably to our knowledge of the molecular mechanism by which insulin stimulates glucose transport.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Physician Scientist Award (K11)
Project #
5K11DK001680-06
Application #
3086334
Study Section
Special Emphasis Panel (ADDK)
Project Start
1989-10-01
Project End
1992-06-30
Budget Start
1991-07-01
Budget End
1992-06-30
Support Year
6
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02115
Sacks, D B; Davis, H W; Crimmins, D L et al. (1992) Casein kinase II-catalysed phosphorylation of calmodulin is altered by amino acid deletions in the central helix of calmodulin. Biochem Biophys Res Commun 188:754-9
Sacks, D B; Davis, H W; Williams, J P et al. (1992) Phosphorylation by casein kinase II alters the biological activity of calmodulin. Biochem J 283 ( Pt 1):21-4
Reddy, G P; Reed, W C; Sheehan, E et al. (1992) Calmodulin-specific monoclonal antibodies inhibit DNA replication in mammalian cells. Biochemistry 31:10426-30
Sacks, D B; McDonald, J M (1992) Effects of cationic polypeptides on the activity, substrate interaction, and autophosphorylation of casein kinase II: a study with calmodulin. Arch Biochem Biophys 299:275-80
Sacks, D B; Davis, H W; Crimmins, D L et al. (1992) Insulin-stimulated phosphorylation of calmodulin. Biochem J 286 ( Pt 1):211-6
Sacks, D B; Porter, S E; Ladenson, J H et al. (1991) Monoclonal antibody to calmodulin: development, characterization, and comparison with polyclonal anti-calmodulin antibodies. Anal Biochem 194:369-77
Sacks, D B; Glenn, K C; McDonald, J M (1989) The carboxyl terminal segment of the c-Ki-ras 2 gene product mediates insulin-stimulated phosphorylation of calmodulin and stimulates insulin-independent autophosphorylation of the insulin receptor. Biochem Biophys Res Commun 161:399-405
Sacks, D B; McDonald, J M (1989) Calmodulin as substrate for insulin-receptor kinase. Phosphorylation by receptors from rat skeletal muscle. Diabetes 38:84-90
Sacks, D B; Fujita-Yamaguchi, Y; Gale, R D et al. (1989) Tyrosine-specific phosphorylation of calmodulin by the insulin receptor kinase purified from human placenta. Biochem J 263:803-12
Fujita-Yamaguchi, Y; Sacks, D B; McDonald, J M et al. (1989) Effect of basic polycations and proteins on purified insulin receptor. Insulin-independent activation of the receptor tyrosine-specific protein kinase by poly(L-lysine). Biochem J 263:813-22

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