The long-range goals of this project include elucidation of the conformational and structure-function relationships of the placental-derived glycoprotein hormone, human chorionic gonadotropin (hCG), delineation of the various intracellular signaling pathways, and characterization of relatively unexplored cellular responses to hCG. A working model has been developed for CG and its pituitary-derived counterpart, luteinizing hormone (LH), that combines the results of chemical and enzymatic modifications, immunological mapping, and physicochemical studies. In order to experimentally test this model, site-directed mutagenesis and trace labeling studies have been designed to elucidate the subunit contact sites for heterodimer formation and the amino acid sequence determinants for receptor binding and activation. Numerous studies have emphasized the role of cAMP in gonadotropin-mediated intracellular signaling, and we also have tentative evidence for the involvement of inositol-1,4,5 trisphosphate and possibly other inositol-- containing phospholipids; this hypothesis of alternate signaling will be experimentally tested in transformed Leydig cells. This project will also delineate some of the cellular responses to gonadotropin and mutant proteins. These include a dramatic CG/LH-mediated morphological change in transformed Leydig cells (MA-10), which we propose is due to cytoskeletal reorganization; a proposed CG/LH-regulated interaction of cells with the extracellular matrix, involving such modulators as SPARC (Secreted Protein that is Acidic and Rich in Cysteine); and CG/LH regulation of the oncogenes c-fos, c-jun, and c-myc. This. comprehensive project is designed to elucidate amino acid sequence determinants regulating heterodimer formation and receptor recognition/activation, intracellular signaling pathways possibly distinct from cAMP, and relatively unexplored aspects of hCG function.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK033973-08A1
Application #
3232383
Study Section
Endocrinology Study Section (END)
Project Start
1983-09-01
Project End
1994-04-30
Budget Start
1991-05-01
Budget End
1992-04-30
Support Year
8
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Miami School of Medicine
Department
Type
Schools of Medicine
DUNS #
City
Miami
State
FL
Country
United States
Zip Code
33146
Chen, Tsuey-Ming; Czerwiec, Frank S; Puett, David (2016) Steroidogenesis and early response gene expression in MA-10 Leydig tumor cells following heterologous receptor down-regulation and cellular desensitization. Biochem Biophys Rep 5:305-312
Cui, Juan; Yin, Yanbin; Ma, Qin et al. (2015) Comprehensive characterization of the genomic alterations in human gastric cancer. Int J Cancer 137:86-95
Bowman, Paula B; Puett, David (2014) Electron paramagnetic resonance spectroscopy of nitroxide-labeled calmodulin. Protein J 33:267-77
Cui, Juan; Xu, Ying; Puett, David (2013) Microarray-based transcriptome profiling of ovarian cancer cells. Methods Mol Biol 1049:119-37
Cui, Juan; Miner, Brooke M; Eldredge, Joanna B et al. (2011) Regulation of gene expression in ovarian cancer cells by luteinizing hormone receptor expression and activation. BMC Cancer 11:280
Angelova, Krassimira; Felline, Angelo; Lee, Moon et al. (2011) Conserved amino acids participate in the structure networks deputed to intramolecular communication in the lutropin receptor. Cell Mol Life Sci 68:1227-39
DeMars, Geneva; Fanelli, Francesca; Puett, David (2011) The extreme C-terminal region of G?s differentially couples to the luteinizing hormone and beta2-adrenergic receptors. Mol Endocrinol 25:1416-30
Cui, Juan; Eldredge, Joanna B; Xu, Ying et al. (2011) MicroRNA expression and regulation in human ovarian carcinoma cells by luteinizing hormone. PLoS One 6:e21730
Puett, David; Angelova, Krassimira; da Costa, Marcelo Rocha et al. (2010) The luteinizing hormone receptor: insights into structure-function relationships and hormone-receptor-mediated changes in gene expression in ovarian cancer cells. Mol Cell Endocrinol 329:47-55
Puett, David; Angelova, Krassimira (2009) Determining the affinity of hormone-receptor interaction. Methods Mol Biol 590:1-20

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