Thyrotropin receptor cleavage at site 1 involves two discontinuous segments at each end of the unique 50-amino acid insertion.
Among the glycoprotein hormone receptors, only the thyrotropin receptor (TSHR) cleaves (at two sites) into disulfide-linked A and B subunits. A 50-amino acid insertion unique to the TSHR ectodomain (residues 317-366) plays no role in ligand binding or signal transduction, but its deletion abrogates cleavage at Site 1, closely upstream of the insertion. We sought to define the region within the 50-amino acid tract involved in TSHR cleavage at Site 1. Mutation of small segments within this region previously failed to prevent cleavage at Site 1. We, therefore, divided the 50-amino acid insertion into quartiles and deleted each one individually (TSHR residues 317-327, 328-338, 339-350, and 351-362). As determined by covalent cross-linking of 125I-TSH to intact cells expressing the mutant receptors, none of these deletions prevented TSHR cleavage at Site 1. Neither did larger deletions of quartiles 1 + 2, 2 + 3, and 3 + 4. However, qualitative differences in the extent of receptor cleavage suggested that quartiles 1 and 4 were playing a greater role in cleavage at Site 1 than were the middle two quartiles. In support of this hypothesis, deletion of these two discontinuous segments almost completely eliminated TSHR cleavage at Site 1. In conclusion, intramolecular cleavage at Site 1 requires the presence of the N-terminal and C-terminal quartiles of the 50-amino acid insertion unique to the TSHR. Taken together with previous observations, our data suggest that this tract may provide a discontinuous binding site for a protease that clips the TSHR at Site 1.
Tanaka
K
K
Autoimmune Disease Unit, Cedars-Sinai Research Institute, Los Angeles, California 90048 and the University of California, Los Angeles School of Medicine, Los Angeles, California 90045, USA.
Chazenbalk
G D
GD
McLachlan
S M
SM
Rapoport
B
B
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009154:Mutation; D011989:Receptors, Thyrotropin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion
D000596:Amino Acids; D011989:Receptors, Thyrotropin