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J. Biol. Chem., Vol. 277, Issue 38, 35013-35018, September 20, 2002

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Identification of Reverb as a Novel ROR Target Gene^*

Philippe Delerive, William W. Chin, and Chen S. Suen

From the Department of Gene Regulation, Bone and Inflammation Research, Eli Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana 46285

The nuclear receptor superfamily comprises a large number of ligand-activated transcription factors that are involved in numerous biological processes such as cell proliferation, differentiation, and homeostasis. ROR (NR1F1) and Reverb (NR1D1) are two members of this family whose biological functions are largely unknown. In addition, no ligand has been yet identified for these two receptors; therefore, they are referred as orphan receptors. Here, we show that ROR and Reverb are expressed with a similar tissue distribution and are both induced during the differentiation of rat L6 myoblastic cells. Ectopic expression of ROR1 in L6 cells significantly induces Reverb expression as demonstrated by Northern blot analysis. Using reverse transcription-PCR to analyze Reverb gene expression from staggerer mice, we found that there was a significant reduction of Reverb mRNA in the skeletal muscle comparing it with the wild-type mice, which suggests that ROR is involved in the regulation of Reverb gene expression. Transient transfection assays using the Reverb promoter demonstrate that ROR regulates the Reverb gene at the transcriptional level. Furthermore, mutagenesis experiments indicate that ROR regulates Reverb transcription via a monomeric ROR response element located in the Reverb gene promoter. Electrophoretic mobility shift assays show that ROR binds strongly to this site in a specific-manner. Finally, overexpression of GRIP-1/TIF-2, but not SRC-1, potentiates ROR-stimulated Reverb promoter activity in transient transfection experiments. Together, our results identify Reverb as a novel target gene for ROR.

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