Differential Recruitment of Nuclear CoRegulators Directs the Isoform-dependent Action of Thyroid Hormone Receptor Mutant

LAURA FOZZATTI; CHANGXUE LU; DONG-WOOK KIM; SHEUE-YANN CHENG

Boston, Massachusetts

Congreso; The 93rd ANNUAL MEETING, ENDO 2011; 2011

The Endocrine Society

Studies using mice deficient in thyroid hormone receptors (TRs) indicate that the two TR isoforms, TRα1 and TRβ1, in addition to mediating overlapping biological activities of the thyroid hormone, T3, also mediate distinct functions. Mice harboring an identical knockin dominant negative mutation (denoted PV) at the C-terminus of TRα1 (Thra1PV mice) or TRβ1 (ThrbPV mice) also exhibit distinct phenotypes. These knockin mice provide an opportunity to understand the molecular basis of isoform-dependent functions in vivo. To test the hypothesis that the distinct functions of TR mutant isoforms are directed by a subset of nuclear regulatory proteins, we first used tandem-affinity chromatography to identify nuclear proteins from HeLa cells that are preferentially associated with TRα1 mutant (TRα1PV) or TRβ1 mutant (TRβ1PV). Thirty-three nuclear proteins including nuclear receptor corepressor (NCoR1) and six other nuclear proteins were identified to preferentially associate with TRα1PV and with TRβ1PV, respectively. Nine common proteins were found to associate with both TR mutants. The involvement of NCoR1 in mediating the distinct phenotype in the liver of Thra1PV and ThrbPV mice was further explored. We showed that NCoR1 preferentially physically interacted with TRα1PV rather than with TRβ1PV in vitro by coimmunoprecipitation assay. The hepatic mRNA levels of CCAAT/enhancer-binding protein α (C/ebpα), a master regulator of adipogenesis, was shown to be down-regulated in the liver of Thra1PV (~50%) but not in ThrbPV mice. A thyroid hormone response element (TRE) was identified in the promoter (-608/-583) of the mouse C/ebpα gene by gel-shift assays. Chromatin immunoprecipitation assay further demonstrated that NCoR1 was preferentially recruited to the mouse TRE-bound TRα1PV on a C/ebpα promoter (2- to 2.5-fold increase vs 0.5-fold on TRE-bound TRβ1PV). This preferential recruitment of NCoR1 by TRα1PV could, at least in part, contribute to the different lipid phenotypic manifestations observed in the liver of these mutant mice. Thus, the present study highlights a novel molecular mechanism by which TR isoforms direct their selective functions via preferential recruitment of a subset of nuclear coregulatory proteins.