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J Biol Chem, Vol. 274, Issue 3, 1394-1400, January 15, 1999

Transcriptional Regulation of the Bmp2 Gene
RETINOIC ACID INDUCTION IN F9 EMBRYONAL CARCINOMA CELLS AND SACCHAROMYCES CEREVISIAE

Loreé C. Heller, Yong Li, Kevin L. Abrams, and Melissa B. Rogers^§¶

From the Departments of  Biology and ^§ Pharmacology and the ^¶ Institute for Biomolecular Science, University of South Florida, Tampa, Florida 33620

Bmp2, a highly conserved member of the transforming growth factor- gene family, is crucial for normal development. Retinoic acid, combined with cAMP analogs, sharply induces the Bmp2 mRNA during the differentiation of F9 embryonal carcinoma cells into parietal endoderm. Retinoic acid (RA) also induces the Bmp2 gene in chick limb buds. Since normal Bmp2 expression may require an endogenous retinoid signal and aberrant Bmp2 expression may cause some aspects of RA-induced teratogenesis, we studied the mechanism underlying the induction of Bmp2. Measurements of the Bmp2 mRNA half-life and nuclear run-on assays indicated that RA stimulated the transcription rate of the Bmp2 gene. The results of ribonuclease protection and primer extension assays indicated that Bmp2 transcription started 2,127 nucleotides upstream of the translation start site in F9 cells. To identify genetic elements controlling this transcription rate increase, upstream and downstream genomic sequences flanking the Bmp2 gene were screened using chloramphenicol acetyltransferase reporter genes in F9 cells and -galactosidase reporter genes in Saccharomyces cerevisiae that were cotransformed with retinoic acid receptor and retinoid X receptor expression plasmids. RA-dependent transcriptional activation was detected between base pairs 2,373 and 2,316 relative to the translation start site. We also identified a required Sp1 binding site between 2,308 and 2,298. The data indicate that Bmp2 is directly regulated by retinoic acid-bound receptors and Sp1.

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