An in vitro model of thyroid neoplasia: permanently transfected FRTL-5 cells with thyroglobulin promoter-cholera toxin A1 subunit minigene.
BACKGROUND
Activating mutations of protein Gs alpha that stimulates adenylyl cyclase are present in a subset of thyroid adenomas. Cholera toxin A1 subunit (CT) mimics these mutations via adenosine diphosphate-ribosylation of Gs alpha.


METHODS
To test the role of activated Gs alpha in thyroid neoplasia we developed an in vitro model. With molecular genetic techniques a transgene (TG-CT) in which the thyroglobulin gene (TG) promoter directs expression of CT was made. This transgene was transfected into rat thyroid follicular cells (FRTL-5). Integration of TG-CT transgene and its expression were examined.


RESULTS
Polymerase chain reaction of DNA from transfected FRTL-5 cells identified the TG-CT transgene in six cell lines. The TG-CT transfected clones exhibited up to a 65-fold (1.29 +/- 0.37 pmols cyclic adenosine monophosphate (cAMP)/micrograms DNA) increase in cAMP over nontransfected FRTL-5 cells (0.02 +/- 0.001 pmols cAMP/micrograms DNA). Insulin, a known stimulator of the TG promoter, further induced CT gene expression and led to a 209-fold (10.43 +/- 0.10 pmols cAMP/micrograms DNA) increase in cAMP over nontransfected cells (0.05 +/- 0.00 pmols cAMP/microgram DNA).


CONCLUSIONS
By mimicking a known mutation associated with thyroid neoplasms, these permanently transfected FRTL-5 cell lines will serve as a model to examine the long-term potential neoplastic effects of activated Gs alpha on thyroid follicular cells.
Saji
M
M
National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Md.
Levine
M A
MA
Zeiger
M A
MA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002772:Cholera Toxin / Q000235:genetics*; D000242:Cyclic AMP / Q000096:biosynthesis; D019204:GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D051381:Rats; D013954:Thyroglobulin / Q000235:genetics*; D013964:Thyroid Neoplasms / Q000209:etiology* / Q000235:genetics; D014162:Transfection
D013954:Thyroglobulin; D002772:Cholera Toxin; D000242:Cyclic AMP; D019204:GTP-Binding Proteins