ISOFORM VARIABLE ACTION AMONG THYROID-HORMONE RECEPTOR MUTANTS PROVIDES INSIGHT INTO PITUITARY RESISTANCE TO THYROID-HORMONE

Resistance to thyroid hormone (RTH) is due to mutations in the beta-is oform of the thyroid hormone receptor (TR-beta). The mutant TR interfe res with the action of normal TR to cause the clinical syndrome. Selec tive pituitary resistance to thyroid hormone (PRTH) results in inappro priate TSH secretion and peripheral sensitivity to elevated thyroid ho rmone levels. Association of the PRTH phenotype with in vitro behavior of the mutant TR has proved elusive. Alternative exon utilization res ults in two TR-beta isoforms, TR-beta 1 and TR-beta 2, which differ on ly in their amino termini. Although the TR-beta 1 isoform is ubiquitou s, the TR-beta 2 isoform is found predominantly in the anterior pituit ary and brain. To date, in vitro evaluation of RTH mutations has focus ed on the TR-beta 1 isoform. Site-directed mutagenesis was used to cre ate several PRTH (R338L, R338W, V349M, R429Q, I431T) and generalized R TH (Delta 337T, P453H) mutations in both TR-beta isoforms. The ability of mutant TRs to act as dominant negative inhibitors of wild type TR- beta function on positive and negative thyroid hormone response elemen ts (pTREs and nTREs, respectively) was evaluated in transient transfec tion assays. PRTH mutants had no significant dominant negative activit y as TR-beta 1 isoforms on pTREs found in peripheral tissues or on nTR Es found on genes regulating TSH synthesis. PRTH mutants, in contrast, had strong dominant negative activity on these same nTREs as TR-beta 2 isoforms. Cotransfected retinoid X receptor-alpha was required for n egative T-3 regulation via the TR-beta 1 isoform but was not necessary for negative regulation via the TR-beta 2 isoform in CV-1 cells. The differing need for retinoid X receptor cotransfection demonstrates two distinct negative T-3-regulatory pathways, one mediated by the TR-bet a 1 and the other mediated by TR-beta 2. The selective effect of PRTH mutations on the TR-beta 2 isoform found in the hypothalamus and pitui tary vs. the TR-beta 1 isoform found in peripheral tissues suggests a molecular mechanism for the PRTH disorder.