Iodothyronine deiodinase activities in FRTL5 cells: predominance of type I 5'-deiodinase.
FRTL5 cells, a thyroid follicular cell line derived from normal rat thyroid, has been extensively used as a model system to study various aspects of the physiology of the thyroid epithelium. The capacity of these cells to metabolize iodothyronines and to generate T3 from T4 has not been previously examined. Here we studied the deiodination of T4, T3, and rT3 in homogenates of FRTL5 cells. By far, these homogenates were more potent catalyzing the 5'-deiodination (outer ring) of T4 and rT3 than the inner ring deiodination of T4 or T3. Both the production of rT3 and the degradation of newly formed T3 from T4 were very limited. Thus, when T4 was used as a substrate, T3 and iodide accumulated in a linear fashion with time, and initially the amounts of iodide and T3 were approximately equal. rT3 and 3,3'-diiodothyronine were rapidly deiodinated by these homogenates, with the 3'-deiodination of 3,3'-diiodothyronine occurring at a slower rate than the 5'-deiodination of rT3. The iodothyronine 5'-deiodinase activity corresponded to type I, as indicated by the following: the Km for T4 and T3 was in the micromolar range; rT3 was a better substrate than T4 (maximum velocity = 101 vs. 19 pmol/min.mg protein; Km = 0.83 vs. 3.1 microM, respectively); and the kinetics of inhibition by 6n-propyl-2-thiouracil were uncompetitive and substrate dependent, suggesting ping-pong kinetics. The type I 5'-deiodinase activity of FRTL5 cells was distinctly stimulated by TSH. This stimulation seems to be mediated by cAMP and requires serum as a permissive factor. In conclusion, 1) FRTL5 cells exhibit both inner and outer ring iodothyronine-deiodinating activities; 2) iodothyronine 5'-deiodination is by far more active; 3) the 5'-deiodination has been characterized as type I deiodinase based on substrate preference, enzyme kinetics, and inhibitors; 4) in all respect iodothyronine deiodination by FRTL5 cell homogenates proceeded with marked similarity to that in homogenates or microsomes of thyroid glands from several species; and 5) the FRTL5 type I deiodinase is more active than that reported in thyroid tissue and as active as that reported in liver and kidney, the prototype of type I deiodinase-containing tissues. The present studies indicate that FRTL5 cells are an excellent model system to study cellular and biochemical aspects of the regulation of this enzyme as well as its regulation by TSH and putative serum factors.
Borges
M
M
Division of Endocrinology, Beth Israel Hospital, Harvard Medical School, Boston, Massachusetts 02215.
Ingbar
S H
SH
Silva
J E
JE
D000818:Animals; D002460:Cell Line; D004229:Dithiothreitol / Q000494:pharmacology; D007453:Iodide Peroxidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007700:Kinetics; D008713:Methimazole / Q000494:pharmacology; D011441:Propylthiouracil / Q000494:pharmacology; D051381:Rats; D013961:Thyroid Gland / Q000187:drug effects / Q000201:enzymology*; D013972:Thyrotropin / Q000494:pharmacology; D013974:Thyroxine / Q000378:metabolism; D014284:Triiodothyronine / Q000378:metabolism; D014285:Triiodothyronine, Reverse / Q000378:metabolism
D014284:Triiodothyronine; D008713:Methimazole; D014285:Triiodothyronine, Reverse; D011441:Propylthiouracil; D013972:Thyrotropin; D007453:Iodide Peroxidase; D013974:Thyroxine; D004229:Dithiothreitol