Thyrotropin Regulation of Malic Enzyme in FRTL-5 Rat Thyroid Cells

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Abstract
TSH-induced increases in malic enzyme mRNA levels in FRTL-5 rat thyroid cells are paralleled by increases in malic enzyme activity and ar mimicked by 8-bromo-cAMP. Apparent approximately 4 h after TSH challenge and maximal after 16 h, they decline by 24 h and are at basal levels by 48 h. The increase occurs in the absence of a measurable effect of TSH on DNA synthesis related to cell growth, since [3H]thymidine incorporation into DNA is still at basal levels 24 h after TSH challenge and is maximal only at 48 h. A protein(s) whose formation is inhibited by cycloheximide appears to be critical to the ability of TSH to increase malic enzyme mRNA levels. Thus, cycloheximide given 30 min before TSH prevents the hormone-induced increase in malic enzyme mRNA; also, when given 24 h after TSH, cycloheximide accelerates the loss of the TSH-induced increase in malic enzyme mRNA levels. A second factor(s) whose formation is prevented by actinomycin-D appears to be important for the decrease in malic enzyme mRNA levels seen 24 and 48 h after TSH challenge. Thus, in experiments in which it is given 24 h after TSH, actinomycin-D preserves the hormone-induced increase in malic enzyme mRNA levels rather than accelerating the decrease, as does cycloheximide. In the same experiment , .beta.-actin mRNA levels decrease to less than 10-20% of control values over the same period; this factor also, therefore, appears to exhibit some degree of specificity. Pulse-chase studies with [3H]uridine indicate that both factors regulate malic enzyme mRNA stability, thus, cycloheximide treatment decreases whereas actinomycin D- increases the apparent half-life of malic enzyme mRNA. The activity of these factors is a means to explain the following observations. First, unlike thyroglobulin, TSH does not increase malic enzymes gene transcription in run-on assays with cell nuclei. Second, neither .alpha.-amanitin nor actinomycin-D given 30 min before TSH inhibits the ability of TSH to induce a relative increase in malic enzyme mRNA levels per unit total RNA measured 6 h after hormone challenge; by comparison, levels of .beta.-actin mRNA are decreased to 50% or less of control values.