Quantitative proteomics of the thyroid hormone receptor-coregulator interactions.
The thyroid hormone receptor regulates a diverse set of genes that control processes from embryonic development to adult homeostasis. Upon binding of thyroid hormone, the thyroid receptor releases corepressor proteins and undergoes a conformational change that allows for the interaction of coactivating proteins necessary for gene transcription. This interaction is mediated by a conserved motif, termed the NR box, found in many coregulators. Recent work has demonstrated that differentially assembled coregulator complexes can elicit specific biological responses. However, the mechanism for the selective assembly of these coregulator complexes has yet to be elucidated. To further understand the principles underlying thyroid receptor-coregulator selectivity, we designed a high-throughput in vitro binding assay to measure the equilibrium affinity of thyroid receptor to a library of potential coregulators in the presence of different ligands including the endogenous thyroid hormone T3, synthetic thyroid receptor beta-selective agonist GC-1, and antagonist NH-3. Using this homogenous method several coregulator NR boxes capable of associating with thyroid receptor at physiologically relevant concentrations were identified including ones found in traditional coactivating proteins such as SRC1, SRC2, TRAP220, TRBP, p300, and ARA70; and those in coregulators known to repress gene activation including RIP140 and DAX-1. In addition, it was discovered that the thyroid receptor-coregulator binding patterns vary with ligand and that this differential binding can be used to predict biological responses. Finally, it is demonstrated that this is a general method that can be applied to other nuclear receptors and can be used to establish rules for nuclear receptor-coregulator selectivity.
Moore
Jamie M R
JM
Department of Pharmaceutical Chemistry, University of California at San Francisco, San Francisco, California 94143-2280, USA.
Galicia
Sarah J
SJ
McReynolds
Andrea C
AC
Nguyen
Ngoc-Ha
NH
Scanlan
Thomas S
TS
Guy
R Kiplin
RK
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D004958:Estradiol / Q000378:metabolism; D047629:Estrogen Receptor beta; D005454:Fluorescence Polarization; D006801:Humans; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D040901:Proteomics / Q000379:methods*; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism; D011987:Receptors, Steroid / Q000235:genetics / Q000378:metabolism; D011988:Receptors, Thyroid Hormone / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D037042:Thyroid Hormone Receptors beta; D014284:Triiodothyronine / Q000037:antagonists & inhibitors
D047629:Estrogen Receptor beta; D019151:Peptide Library; D010455:Peptides; D011960:Receptors, Estrogen; D011987:Receptors, Steroid; D011988:Receptors, Thyroid Hormone; D037042:Thyroid Hormone Receptors beta; D014284:Triiodothyronine; D004958:Estradiol