Isolation and characterization of a zebrafish homologue of the cone rod homeobox gene.
OBJECTIVE
To isolate and characterize a zebrafish CRX: homologue. Mammalian CRX: genes are expressed specifically in photoreceptors and pinealocytes, regulate photoreceptor gene expression, are necessary for normal photoreceptor differentiation, and when mutated cause a variety of photoreceptor degenerations.


METHODS
A zebrafish retinal cDNA library was screened with a human CRX cDNA probe. Radiation hybrid mapping, Northern blot analysis, in situ hybridization, and transient transfection studies were performed using standard methods.


RESULTS
Based on amino acid sequence comparisons, zebrafish crx shows 50% identity with human CRX, and 85% identity in the homeodomain. A phylogenetic analysis indicates that zebrafish crx is most closely related to the mammalian Crx proteins, and more distantly related to the Otx proteins. Zebrafish crx maps between 49.6 and 54.5 cm from the top of linkage group LG05C, a map position consistent with the location of the mouse and human CRX genes. Northern blot analysis and in situ hybridization indicate that zebrafish crx is expressed in the retina and pineal gland. In adult zebrafish, crx is expressed by both rods and cones in the outer nuclear layer, and in cells in the outer zone of the inner nuclear layer, in the region occupied by bipolar cells. Similar to mammalian Crx, zebrafish crx interacts with neural retinal leucine zipper (Nrl) to activate, although weakly, rhodopsin promoter activity.


CONCLUSIONS
Based on molecular phylogeny, chromosomal location, expression pattern, and ability to activate rhodopsin promoter activity in transient transfection assays, zebrafish crx appears to be an orthologue and functional homologue of mammalian CRX:
Liu
Y
Y
Department of Ophthalmology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Shen
Y
Y
Rest
J S
JS
Raymond
P A
PA
Zack
D J
DJ
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004247:DNA / Q000032:analysis; D015342:DNA Probes; D015723:Gene Library; D008040:Genetic Linkage; D018398:Homeodomain Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification*; D006801:Humans; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D051857:Otx Transcription Factors; D010870:Pineal Gland / Q000737:chemistry; D012160:Retina / Q000737:chemistry* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification*; D014162:Transfection; D015027:Zebrafish
D015342:DNA Probes; D018398:Homeodomain Proteins; D051857:Otx Transcription Factors; D015534:Trans-Activators; C109288:cone rod homeobox protein; D004247:DNA