Characterization of the thyroid hormone response element in the skeletal alpha-actin gene: negative regulation of T3 receptor binding by the retinoid X receptor.
We have identified a T3 response element (TRE) in the human skeletal alpha-actin gene between nucleotide positions -273 and -249 (5' GGGCAACTGGGTCGGGTCAGGAGGG 3') that is accommodated by the core receptor binding motif, A/G GG T/A C A/G. This sequence conferred appropriate hormonal regulation in a thyroid hormone receptor (TR alpha) dependent manner to an enhancerless SV40 promoter. Electrophoretic mobility shift assay experiments showed that Escherichia coli expressed and affinity purified TR alpha bound to the skeletal alpha-actin TRE in a sequence specific manner. The alpha-actin TRE bound TR alpha dimers cooperatively. Mutagenesis of the alpha-actin TRE indicated that the core binding motifs and the gap sequences were the most important for efficient binding to TR alpha. The retinoid X receptor alpha (RXR alpha) interacted with the alpha-actin TRE in a sequence specific fashion and formed heterodimeric complexes with TR alpha on the alpha-actin TRE. However, increased levels of RXR alpha decreased the binding of TR alpha to the alpha-actin TRE, in contrast to promoting TR alpha binding to the alpha-myosin heavy chain TRE. Furthermore, the alpha-actin, palindromic, synthetic direct repeat, alpha-myosin heavy chain, and growth hormone TREs interacted with an identical nuclear factor in vitro in muscle cells. In conclusion, our data suggest that the human skeletal alpha-actin TRE is a target for direct cross-talk between two different hormonal signals (T3 and 9-cis-retinoic acid) at the receptor level.(ABSTRACT TRUNCATED AT 250 WORDS)
University of Queensland, Centre for Molecular Biology and Biotechnology, Ritchie Research Laboratories, St. Lucia, Australia.
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Muscat
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Griggs
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Downes
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Emery
D000199:Actins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D001842:Bone and Bones / Q000737:chemistry*; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011956:Receptors, Cell Surface / Q000502:physiology; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D047488:Retinoid X Receptors; D013539:Simian virus 40 / Q000235:genetics; D014157:Transcription Factors; D014284:Triiodothyronine / Q000502:physiology*
D000199:Actins; D011956:Receptors, Cell Surface; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone; D011994:Recombinant Proteins; D047488:Retinoid X Receptors; D014157:Transcription Factors; D014284:Triiodothyronine