Single subunit structure of the human thyrotropin receptor

Area of Science:

• Endocrinology
• Molecular Biology
• Immunology

Background:

• The structure of the human thyrotropin receptor (hTSH-R) has been debated, with some models suggesting a two-subunit composition.
• Understanding the receptor's subunit structure is crucial for comprehending its function in thyroid hormone regulation.

Purpose of the Study:

• To generate a specific antibody against the N-terminal extracellular domain of hTSH-R.
• To elucidate the subunit composition of the hTSH-R using the developed antibody.

Main Methods:

• Production of a rabbit polyclonal antibody against a synthetic peptide (residues 29-57) from the hTSH-R N-terminus.
• Western blot analysis of recombinant hTSH-R in CHO-K1 cells and rat TSH-R in FRTL5 cells.
• Antibody specificity was confirmed through absorption with the N-peptide.

Main Results:

• The N-peptide antibody recognized recombinant hTSH-R at approximately 104 kDa under both reducing and non-reducing conditions.
• The antibody did not bind to untransfected cells or when pre-absorbed with the N-peptide, confirming specificity.
• The antibody also detected rat TSH-R at a similar molecular weight (104 kDa) under reducing conditions.

Conclusions:

• The findings strongly support that the TSH-R functions as a single subunit.
• The previously proposed two-subunit model for TSH-R likely resulted from artifactual receptor cleavage during sample preparation.