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      Cloning and functional expression of a thyrotropin receptor cDNA from rat fat cells.

      The Journal of Biological Chemistry
      Adipose Tissue, metabolism, Animals, Base Sequence, DNA Primers, chemistry, DNA, Complementary, genetics, Epididymis, Gene Expression, Lipid Mobilization, drug effects, Male, Molecular Sequence Data, RNA, Messenger, Rats, Receptors, Thyrotropin, Sequence Alignment, Sequence Homology, Nucleic Acid, Thyroid Gland, Thyrotropin, pharmacology

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          Abstract

          Thyrotropin receptor (TSH-R) has been thought to be thyroid-specific, but, by Northern blot analysis, we found that rat adipose tissue expressed TSH-R mRNAs in amounts approaching those in the thyroid. To investigate the function of TSH-R from adipose tissue, we screened a rat fat cell lambda gt11 cDNA library for TSH-R sequences using a 32P-labeled rat thyroid TSH-R cDNA as a probe. Among 10(6) plaques, we obtained four positive clones. Sequencing of these cDNAs has revealed that two of them (F alpha and F beta) contained both initiation and termination codons. Comparison of F alpha with the thyroid TSH-R cDNA sequence revealed that F alpha was almost identical to the thyroid TSH-R, except that nucleotides 1041 and 1277 were changed from A to G and from C to T, respectively. In contrast, we found that F beta contained 21 novel nucleotides between nucleotides 467 and 468 of the thyroid TSH-R cDNA, encoding an additional 7 amino acids. However, when we prepared mRNA from adipose tissue and transcribed it into cDNA, we failed to amplify the F beta type of TSH-R cDNA by polymerase chain reaction, suggesting that F beta mRNAs are rare in the tissue. We then ligated F cDNAs into pSG5 and transfected them with pSV2-neo into Chinese hamster ovary (CHO)-K1 cells. TSH stimulated cAMP formation in CHO-F alpha cells in a manner similar to that in CHO cells transfected with thyroid TSH-R cDNA. In contrast, no increase of cAMP was observed in CHO-F beta cells. IgG from patients with Graves' disease (n = 4) showed thyroid-stimulating antibody activity only in CHO-F alpha cells (1288-4582%). In addition, CHO-F alpha cells and CHO cells transfected with thyroid TSH-R showed similar 125I-TSH binding activity. These results indicate that the fat cell expresses high levels of a TSH-R whose function is indistinguishable from that in the thyroid and suggest that the TSH-R autoantibody plays an important role in the pathogenesis of the extrathyroidal manifestations of Graves' disease.

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