Signal transduction by the human thyrotropin receptor: studies on the role of individual amino acid residues in the carboxyl terminal region of the third cytoplasmic loop.
We observed previously that the carboxyl-terminal region of the third loop of the TSH receptor (amino acid residues 617-625) is important in signal transduction. To analyze this region in more detail, in the present study we used site-directed mutagenesis to substitute, on an individual basis, the seven amino acids previously mutated as a group. These amino acids are either charged residues or potential phosphorylation sites. Six of the mutant TSH receptors with individual amino acid substitutions bound TSH with high affinity and displayed a cAMP response to TSH stimulation similar to the wild-type TSH receptor. The mutant receptor TSH-R-Gly625 (Arg----Gly) did not transduce a signal, but these results are noninformative because of the loss of high affinity TSH binding. The present data indicate that for each of the six informative amino acid substitutions, the individual residues are not critical for signal transduction. A corollary of this conclusion is that in the important carboxyl-terminal region of the third cytoplasmic loop of the TSH receptor multiple amino acid residues function as a unit.
Chazenbalk
G D
GD
Thyroid Molecular Biology Unit, Veterans Administration Medical Center, San Francisco, California.
Nagayama
Y
Y
Wadsworth
H
H
Russo
D
D
Rapoport
B
B
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D008961:Models, Structural; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011989:Receptors, Thyrotropin / Q000235:genetics / Q000502:physiology*; D015183:Restriction Mapping; D015398:Signal Transduction; D013972:Thyrotropin / Q000378:metabolism* / Q000494:pharmacology; D014162:Transfection
D011989:Receptors, Thyrotropin; D013972:Thyrotropin; D000242:Cyclic AMP