Functional regulation of thyroid hormone receptor variant TR alpha 2 by phosphorylation.
The thyroid hormone (T3) receptor (TR) variant TR alpha 2 is abundant in brain but does not bind T3 because of its unique C terminus. The only known function of TR alpha 2, inhibition of TR-dependent transactivation, involves competition for T3 response elements. Paradoxically, in vitro-translated TR alpha 2 bound poorly to these sites. We report here that dephosphorylation of TR alpha 2 restored its DNA binding. Mutation of C-terminal serine residues to alanine (TR alpha 2-SA) was equally effective. The C terminus of TR alpha 2 was phosphorylated in a human cell line, whereas that of TR alpha 2-SA was not. Conversely, TR alpha 2-SA was a much better inhibitor of T3 action than was wild-type TR alpha 2. The dominant negative activity of TR alpha 2-SA was less than stoichiometric with TR concentration, possibly because it was unable to heterodimerize with retinoid X receptor, which enhances the binding of other TRs. Purified casein kinase II as well as a reticulocyte casein kinase II-like activity phosphorylated TR alpha 2 on serines 474 and 475. Mutation of these two residues to alanine was sufficient to restore DNA binding. Thus, DNA binding by TR alpha 2 is regulated by phosphorylation at a site distant from the DNA-binding domain. The increased dominant negative activity of a nonphosphorylatable form of TR alpha 2 suggests that phosphorylation may provide a rapid, T3-independent mechanism for cell-specific modulation of the expression of T3-responsive genes.
Katz
D
D
Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Reginato
M J
MJ
Lazar
M A
MA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D047390:Casein Kinase II; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D014644:Genetic Variation; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017354:Point Mutation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D011988:Receptors, Thyroid Hormone / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection
D017931:DNA Primers; D010750:Phosphoproteins; D011988:Receptors, Thyroid Hormone; D004247:DNA; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases