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J. Biol. Chem., Vol. 276, Issue 18, 15146-15154, May 4, 2001
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From the Department of Biology, National Science Foundation Center
for Biological Timing, University of Virginia,
Charlottesville, Virginia 22904-4328
Nocturnin is a vertebrate circadian
clock-regulated gene, and in Xenopus laevis its mRNA is
specifically expressed in retinal photoreceptor cells. We have
investigated the transcriptional regulatory mechanism that drives this
precise spatial expression pattern of the nocturnin gene. A deletion
series of the nocturnin 5'-flanking sequence driving the green
fluorescence protein (GFP) reporter was used to generate transgenic
Xenopus tadpoles. We found that a construct containing 2.6 kilobase pairs of 5'-flanking sequence targeted high level GFP reporter
expression specifically to photoreceptor cells, in a pattern identical
to endogenous nocturnin. This photoreceptor-specific expression pattern
was maintained with several further deletions of 5'-upstream sequence,
including a short 59-base pair fragment. Within this region of 59 base
pairs, three perfect repeats of a novel protein binding site were
identified by electrophoretic mobility shift assay. Competitions
using varying oligonucleotide sequences demonstrated that the sequence
required for protein binding is CAGACAGGCTATA, designated
photoreceptor-conserved element II (PCE II). The protein complex that
binds to this element is enriched in retinal extracts, and mutations of
PCE II which fail to bind the protein complex also fail to direct GFP
reporter expression to photoreceptors. These results indicate that the PCE II in the proximal promoter of the nocturnin gene is sufficient for
driving the photoreceptor-specific expression of nocturnin.
A Novel Promoter Element, Photoreceptor Conserved
Element II, Directs Photoreceptor-specific Expression of
Nocturnin in Xenopus laevis*
*
This work was funded by Grant EY11489 from the NEI, National
Institutes of Health (to C. B. G.) and by the Karl Kirchgessner Foundation (to C. B. G.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: 264 Gilmer Hall, Dept.
of Biology, University of Virginia, Charlottesville, VA 22904-4328. Tel.: 804-982-5436; Fax: 804-982-5626; Email:
cbg8b@virginia.edu.
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