The primary objective of this grant proposal is to further our understanding of the TSH receptor, in terms of its physiological interaction with TSH and its pathophysiological role in autoimmune thyroid disease. The following studies will be performed: 1. Studies on TSH binding to the TSH receptor: The binding of TSH to recombinant human TSH receptor variants will be used as a means to understand the structure of the receptor, in particular to help localize the specific regions that interact with TSH. Approaches to be used include the use of: a) TSH-LH/CG receptor chimeras, and b) individual amino acid substitutions. 2. TSH receptor signal transduction: TSH-LH/CG receptor chimeras as well as other TSH receptor mutants will be used to identify the extracellular domains and cytoplasmic regions of the TSH receptor that are involved in signal transduction. The role of these regions in both the CAMP and phosphatidyl inositol pathways will be studied. A possible role for phosphorylation in TSH receptor function will be investigated. 3. Overexpression and crystallization of the TSH receptor: Large quantities of recombinant TSH receptor protein are a requirement for crystallization and determination of the 3-dimensional structure of the molecule. Purified TSH receptor protein will also be invaluable for the generation of mouse and human TSH receptor monoclonal antibodies. Two approaches will be undertaken to attain this goal; namely dihydrofolate reductase (DHFR)-linked amplification and baculovirus expression. 4. Study of TSH receptor antibodies: The primary goal will be to define the epitopes on the TSH receptor that are recognized by TSH receptor antibodies in the sera of patients with autoimmune thyroid disease. Secondary goals will be the generation of monoclonal antibodies for TSH receptor purification. The approaches to be taken include: a) Screening of a TSH receptor CDNA fragment expression library with sera from patients with autoimmune thyroid disease; b) Generation of mouse anti-TSH receptor monoclonal antibodies; c) Generation of human anti-TSH receptor monoclonal antibodies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK019289-19
Application #
2137289
Study Section
Endocrinology Study Section (END)
Project Start
1976-05-01
Project End
1997-04-30
Budget Start
1994-05-18
Budget End
1995-04-30
Support Year
19
Fiscal Year
1994
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Rapoport, Basil; McLachlan, Sandra M (2016) TSH Receptor Cleavage Into Subunits and Shedding of the A-Subunit; A Molecular and Clinical Perspective. Endocr Rev 37:114-34
Rapoport, Basil; Banuelos, Bianca; Aliesky, Holly A et al. (2016) Critical Differences between Induced and Spontaneous Mouse Models of Graves' Disease with Implications for Antigen-Specific Immunotherapy in Humans. J Immunol 197:4560-4568
Chen, Chun-Rong; Hubbard, Paul A; Salazar, Larry M et al. (2015) Crystal structure of a TSH receptor monoclonal antibody: insight into Graves' disease pathogenesis. Mol Endocrinol 29:99-107
Chen, Chun-Rong; Salazar, Larry M; McLachlan, Sandra M et al. (2015) Deleting the Redundant TSH Receptor C-Peptide Region Permits Generation of the Conformationally Intact Extracellular Domain by Insect Cells. Endocrinology 156:2732-8
Rapoport, Basil; Aliesky, Holly A; Banuelos, Bianca et al. (2015) A unique mouse strain that develops spontaneous, iodine-accelerated, pathogenic antibodies to the human thyrotrophin receptor. J Immunol 194:4154-61
Rapoport, Basil; Aliesky, Holly A; Chen, Chun-Rong et al. (2015) Evidence that TSH Receptor A-Subunit Multimers, Not Monomers, Drive Antibody Affinity Maturation in Graves' Disease. J Clin Endocrinol Metab 100:E871-5
Rapoport, Basil; McLachlan, Sandra M (2014) Graves' hyperthyroidism is antibody-mediated but is predominantly a Th1-type cytokine disease. J Clin Endocrinol Metab 99:4060-1
McLachlan, Sandra M; Rapoport, Basil (2014) Breaking tolerance to thyroid antigens: changing concepts in thyroid autoimmunity. Endocr Rev 35:59-105
Aliesky, Holly; Courtney, Cynthia L; Rapoport, Basil et al. (2013) Thyroid autoantibodies are rare in nonhuman great apes and hypothyroidism cannot be attributed to thyroid autoimmunity. Endocrinology 154:4896-907
McLachlan, Sandra M; Rapoport, Basil (2013) Thyrotropin-blocking autoantibodies and thyroid-stimulating autoantibodies: potential mechanisms involved in the pendulum swinging from hypothyroidism to hyperthyroidism or vice versa. Thyroid 23:14-24

Showing the most recent 10 out of 136 publications