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(Received for publication, November 6, 1995; and in revised form, January 11, 1996) A previously characterized retinoic acid response element
(RARE1) in the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter
confers approximately 50% of the response of this gene to retinoic acid
(RA). Transient transfection experiments were performed using
constructs containing progressive 5` deletions of the PEPCK promoter to
locate other elements that contribute to the RA response. A second RARE
(RARE2) was located between -402 and -306. Methylation
interference and mobility gel shift assays indicated that RAR/RXR bound
specifically to a segment of DNA located between -337 and
-321. This region contains consensus and degenerate half-sites
for receptor binding separated by 5 bp. Mutations in either half-site
selectively decreased the RA response and diminished RAR/RXR binding in
mobility gel shift assays. When both RARE1 and RARE2 were mutated, 80%
of the RA response was lost. Finally, RARE2 conferred a RA response in
a heterologous promoter context. We conclude that RAR/RXR binds to
RARE2, and that this DR5-type element is a major contributor to the
response of the PEPCK gene to RA.
Volume 271,
Number 11,
Issue of March 15, 1996 pp. 6260-6264
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
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