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J. Biol. Chem., Vol. 275, Issue 38, 29794-29799, September 22, 2000
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§,§,
,**§§, and¶¶
From the Departments of Photoreceptor-specific expression of rhodopsin is
mediated by multiple cis-acting elements in the proximal promoter
region. NRL (neural retina leucine
zipper) and CRX (cone rod homeobox) proteins bind to the adjacent NRE and Ret-4 sites, respectively, within
this region. Although NRL and CRX are each individually able to induce
rhodopsin promoter activity, when expressed together they exhibit
transcriptional synergy in rhodopsin promoter activation. Using the
yeast two-hybrid method and glutathione S-transferase pull-down assays, we demonstrate that the leucine zipper of NRL can
physically interact with CRX. Deletion analysis revealed that the CRX
homeodomain (CRX-HD) plays an important role in the interaction with
the NRL leucine zipper. Although binding with the CRX-HD alone was
weak, a strong interaction was detected when flanking regions including
the glutamine-rich and the basic regions that follow the HD were
included. A reciprocal deletion analysis showed that the leucine zipper
of NRL is required for interaction with CRX-HD. Two disease-causing
mutations in CRX-HD (R41W and R90W) that exhibit reduced DNA binding
and transcriptional synergy also decrease its interaction with NRL.
These studies suggest novel possibilities for protein-protein
interaction between two conserved DNA-binding motifs and imply that
cross-talk among distinct regulatory pathways contributes to the
establishment and maintenance of photoreceptor function.
Ophthalmology and Visual
Sciences and ¶¶ Human Genetics, W. K. Kellogg Eye
Center, University of Michigan, Ann Arbor, Michigan 48105, the
¶ Department of Ophthalmology and Visual Sciences,
Washington University School of Medicine, St. Louis, Missouri
63110, and the Departments of ** Ophthalmology,
Molecular Biology and Genetics, and
§§ Neuroscience, The Johns Hopkins University
School of Medicine, Baltimore, Maryland 21287
Recipient of an RPB Career Development Award.
Recipient of a Lew R. Wasserman Merit Award from RPB.
To whom correspondence should be addressed: Dept. of Ophthalmology and Visual Sciences, Kellogg Eye Center, University of Michigan, 1000 Wall
St., Ann Arbor, MI 48105. Tel.: 734-763-3731; Fax: 734-647-0228; E-mail: swaroop@umich.edu.
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