Abstract
Recent studies from this laboratory have demonstrated the presence of thyroid hormone response elements (TREs) in the 5'-flanking region of the rat alpha and TSH beta subunit genes. Using an avidin-biotin complex DNA binding assay, we have shown that these TREs bind the thyroid hormone (T3) receptor present in nuclear extracts of GH3 cells, as well as the in vitro synthesized Hc-erbA beta, which has been identified as a member of the family of T3 receptors. The binding of Hc-erbA beta to the alpha subunit TRE can be enhanced 3-4-fold by including GH3 nuclear extract in the binding assay. Binding to the TRE present in the TSH beta gene or the rat growth hormone gene was similarly enhanced, although to a lesser degree. The enhanced binding activity is trypsin-sensitive and heat labile, and is not reproduced by the addition of histones, bovine serum albumin, or cytosol instead of nuclear extract. Gel exclusion chromatography suggests a molecular size of approximately 65,000 Da. This protein, which is present in several different cell types, is also able to complement binding of the rat erbA alpha-1 and the pituitary-specific erbA beta-2 forms of the receptor. These data suggest that the binding of the T3 receptor to a TRE is augmented by another nuclear protein, which may be involved in the mechanism of action of thyroid hormone.
Highlights
From the Division of Genetics, Department of Medicine, Brigham and Women’s Hospital, Howard Hughes Medical Institute and Harvard Medical Sihool, Boston, kassachukts
The addition of 50 c(g of nuclear extract to the binding reaction increased receptor binding to 300% ((Y subunit), 150% (TSHP) or 160% of the binding observed in the absence of nuclear extract
The addition of increasing amounts of nuclear extract to the binding reaction increased the amount of Hc-erbA/3 bound to the (Y subunit gene sequences (Fig. 2) until a plateau was reached
Summary
From the Division of Genetics, Department of Medicine, Brigham and Women’s Hospital, Howard Hughes Medical Institute and Harvard Medical Sihool, Boston, kassachukts Recent studies from this laboratory have demonstrated the presence of thyroid hormone response elements (TREs) in the 5’-flanking region of the rat This protein, which is present in several different cell types, is able to complement binding of the rat erbAa-1 and the pituitaryspecific erbA&2 forms of the receptor. These data suggest that the binding of the Ts receptor to a TRE is augmented by another nuclear protein, which may be involved in the mechanism of action of thyroid hormone
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