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Volume 271, Number 44, Issue of November 1, 1996 pp. 27919-27926
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

Negative Regulation of the Gene for the Preprothyrotropin-releasing Hormone from the Mouse by Thyroid Hormone Requires Additional Factors in Conjunction with Thyroid Hormone Receptors

(Received for publication, June 5, 1996, and in revised form, July 30, 1996)

Teturou Satoh , Masanobu Yamada , Toshiharu Iwasaki and Masatomo Mori

From the First Department of Internal Medicine, Gunma University School of Medicine, 3-39-15 Showa-machi, Maebashi 371, Japan

To gain additional insights into the negative gene regulatory action by triiodothyronine (T3), we isolated a 2-kilobase pair 5'-flanking region of the mouse preprothyrotropin-releasing hormone (ppTRH) gene and characterized the DNA elements mediating inhibitory regulation by T3 in the promoter region. In GH4C1 cells, the expression of the 2-kilobase pair mouse ppTRH 5'-flanking region fused to the luciferase reporter gene occurred by transfection and was significantly suppressed by T3. In contrast, T3 suppression was not observed in T3 receptor (T3R)-deficient CV-1 cells, suggesting that T3Rs were required for the negative regulation. Cotransfected mouse T3R alpha 1, beta 1, and beta 2 possessed indistinguishable potency for the negative regulation. Deletion analysis localized the element mediating the negative regulation to the region between -83 and +46, and the sequence downstream of the transcription start site (TSS) between +12 and +46 was found to be essential for the inhibitory regulation. In mobility shift assays, only T3R monomers bound to the element containing a T3 response element half-site at -57. No apparent T3R binding was observed to the element downstream of TSS. Neither the T3 response element half-site nor the element downstream of the TSS confer T3 suppression individually in heterologous promoters. These results indicate that the negative regulation of murine ppTRH gene by T3 might be mediated by the cooperation of T3R monomers with unknown factor(s) interacting with the element downstream of the TSS.


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