Transfection with the cDNA of the human thyrotropin receptor of a poorly differentiated rat thyroid cell line (FRT).
A cell line derived from the Fisher rat thyroid (FRT), that does not have functional TSH receptor, was stably transfected with the cDNA of the human TSH receptor (h TSH-R). In wild FRT cells TSH (1-1000 mU/l) was unable to increase cAMP production, while 10-10000 nmol/l forskolin elicited a 10-30 fold cAMP stimulation. Two of the transfected clones were responsive to TSH in terms of cAMP production. In particular, the FRT-R3 transfected clone showed the highest sensitivity to the hormone with a 10 fold cAMP increase over the basal at 100 mU/l TSH. The Northern blot analysis using a 2.4 kbp cDNA probe for the hTSH-R showed a band corresponding to the mRNA of TSH receptor in FRT-R3 cells, but not in wild FRT cells. In both cell types TSH was ineffective in stimulating growth assayed by 3H-thymidine incorporation into DNA. Hybridization with a probe for thyroperoxidase on polymerase chain reaction products after reverse transcription of mRNA showed that FRT-R3, as well as FRT cells, do not have a transcript for thyroperoxidase. In conclusion, the data reported in this paper show that the insertion of the hTSH-R cDNA in the genome of poorly differentiated rat thyroid cells results in the recovery of TSH-dependent adenylate cyclase, but not other differentiated thyroid cell functions.
Elisei
R
R
Istituto di Endocrinologia, University of Pisa, Italy.
Pinchera
A
A
Chiovato
L
L
Mammoli
C
C
Agretti
P
P
Romei
C
C
Santini
F
F
Bendinelli
G
G
Fiore
E
E
Capaccioli
A
A
Vitti
P
P
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D018076:DNA, Complementary / Q000235:genetics*; D016678:Genome; D006801:Humans; D051381:Rats; D011916:Rats, Inbred F344; D011989:Receptors, Thyrotropin / Q000235:genetics*; D013961:Thyroid Gland / Q000166:cytology*; D014162:Transfection
D018076:DNA, Complementary; D011989:Receptors, Thyrotropin; D000262:Adenylyl Cyclases