This is a proposal to investigate some aspects of iodine and thyroid hormone metabolism and will focus on (I) the mechanism(s) and biochemical and physiological regulation of thyroid hormone deiodination in rat tissues, with emphasis on the formation of T3 from T4, (II) identification and characterization of the thiol cofactor mechanism responsible for the activation of thyroid hormone deiodination in vivo, (III) roles of cAMP and cGMP in the espression of thyroid hormone action at the cellular level, and (IV) characterization of a thyroaidal electron-transfer system in relation to the NADPH-dependent deiodination of iodotyrosines. Goal (I) will be approached by attempts at solubilization, purification and kinetic characterization of the 'high' and 'low' Km iodothyronine 5'-deiodinases in various tissues as well as by in vivo studies aimed at assessing their relative contribution to circulating T3 in various thyroid states. Goal (II) will characterize (a) the cytosolic thiol protein that promotes GSH-activation of iodothyronine 5'-deiodinases in vitro, and examine its relationship to glutaredoxin, and (b) another soluble thiol protein which catalyzes oxidative degradation of iodothyronines and evaluate its possible role as a disposal mechanism for excess iodothyronines. Goal (III) will examine the effects of thyroid status on the enzymatic properties, molecular forms and dynamic interrelationships of cAMP- and cGMP-phosphodiesterases in adipocytes and their concerted effects on the steady state levels of cellular cAMP and cGMP. Goal (IV) will be pursued experimentally by purification of thyroid ferredoxin and NADPH-ferredoxin reductase, establishment of its role in deiodination of iodotyrosines and characterization of the enzymatic and molecular properties of the reductase enzyme in relation to protease-purified preparations of thyroid iodotyrosine deiodinase.
