The adipocyte hormone leptin regulates energy balance, substrate metabolism, immunity, bone formation and reproduction. Our recent work demonstrates that leptin production is regulated at the translational level, and that elements within its 5' UTR stimulate and its 3' UTRs inhibit translation of a reporter gene. The major goal of this application is to identify the cis elements and trans acting factors that regulate leptin translation in response to variations in nutritional state. The objectives of this application are: 1) To identify RNA binding proteins (RBP) that interact with the leptin 5' and 3'UTRs, and define the cis elements involved; 2) To determine the effect of starvation/feeding in vivo and acute treatment with insulin and beta-adrenergic agonists in vitro on expression of RBPs and their interaction with leptin mRNA; 3) To assess the functional roles of specific RBPs in leptin mRNA translation using knockdown and overexpression studies; and 4) To delineate the signaling mechanisms that cause high basal and insulin-resistant leptin translation in obesity. The leptin 3'UTR includes AU-rich sequences and motifs for binding of RBPs that are known to stimulate (HuR) or inhibit (TIA-1, TIAR) the translation of specific mRNAs. We will therefore determine if leptin mRNA 'in vivo' associates with RBPs 'in vivo' by immunoprecipitating ribonucleoprotein complexes from cytosolic extracts of 3T3-L1 adipocytes and rat adipocytes. Pull-down assays using biotinylated leptin UTR probes will verify the interactions of RBPs of interest and point to the motifs involved. We will test the hypothesis that insulin in vitro or nutritional status in vivo (starvation, feeding) affects the binding of specific RBPs to leptin mRNA, and involves an alteration in their expression and/or nucleocytoplasmic shuttling. Finally, we will test the hypothesis that the chronic hyperinsulinemia associated with obesity increases leptin production at the translational level through the activation of stress / energy sensing pathways (e.g. mTOR, AMPK, and MAPK) and/or by altering the expression or binding of specific RBPs to leptin mRNA. Overall, the proposed studies will enhance understanding of leptin biology and broaden knowledge of how the adipocyte functions as an endocrine cell that orchestrates the production of numerous hormones and cytokines in response to nutritional cues. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK052398-10
Application #
7477183
Study Section
Cellular Aspects of Diabetes and Obesity Study Section (CADO)
Program Officer
Haft, Carol R
Project Start
1997-09-06
Project End
2008-12-31
Budget Start
2008-07-01
Budget End
2008-12-31
Support Year
10
Fiscal Year
2008
Total Cost
$95,511
Indirect Cost
Name
University of Maryland Baltimore
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
188435911
City
Baltimore
State
MD
Country
United States
Zip Code
21201
Lee, M-J; Fried, S K (2014) The glucocorticoid receptor, not the mineralocorticoid receptor, plays the dominant role in adipogenesis and adipokine production in human adipocytes. Int J Obes (Lond) 38:1228-33
Lee, Mi-Jeong; Pramyothin, Pornpoj; Karastergiou, Kalypso et al. (2014) Deconstructing the roles of glucocorticoids in adipose tissue biology and the development of central obesity. Biochim Biophys Acta 1842:473-81
Lee, Mi-Jeong; Fried, Susan K (2014) Optimal protocol for the differentiation and metabolic analysis of human adipose stromal cells. Methods Enzymol 538:49-65
Lee, Mi-Jeong; Wu, Yuanyuan; Fried, Susan K (2013) Adipose tissue heterogeneity: implication of depot differences in adipose tissue for obesity complications. Mol Aspects Med 34:1-11
Carswell, Kirstin A; Lee, Mi-Jeong; Fried, Susan K (2012) Culture of isolated human adipocytes and isolated adipose tissue. Methods Mol Biol 806:203-14
Lee, Mi-Jeong; Fried, Susan K (2012) Glucocorticoids antagonize tumor necrosis factor-?-stimulated lipolysis and resistance to the antilipolytic effect of insulin in human adipocytes. Am J Physiol Endocrinol Metab 303:E1126-33
Nimitphong, Hataikarn; Holick, Michael F; Fried, Susan K et al. (2012) 25-hydroxyvitamin D? and 1,25-dihydroxyvitamin D? promote the differentiation of human subcutaneous preadipocytes. PLoS One 7:e52171
Lee, Mi-Jeong; Wu, Yuanyuan; Fried, Susan K (2012) A modified protocol to maximize differentiation of human preadipocytes and improve metabolic phenotypes. Obesity (Silver Spring) 20:2334-40
Lee, Eun Kyung; Lee, Mi Jeong; Abdelmohsen, Kotb et al. (2011) miR-130 suppresses adipogenesis by inhibiting peroxisome proliferator-activated receptor gamma expression. Mol Cell Biol 31:626-38
Lee, Mi-Jeong; Gong, Da-Wei; Burkey, Bryan F et al. (2011) Pathways regulated by glucocorticoids in omental and subcutaneous human adipose tissues: a microarray study. Am J Physiol Endocrinol Metab 300:E571-80

Showing the most recent 10 out of 27 publications