IOVS Arteriosclerosis, Thrombosis, and Vascular Biology
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(Investigative Ophthalmology and Visual Science. 2006;47:4245-4253.)
© 2006 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.06-0167
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The Rx-like Homeobox Gene (Rx-L) Is Necessary for Normal Photoreceptor Development

Yi Pan,1 Srivamsi Nekkalapudi,1 Lisa E. Kelly,1 and Heithem M. El-Hodiri1,2

1From the Center for Molecular and Human Genetics, Columbus Children’s Research Institute, and the 2Department of Pediatrics, School of Medicine and Public Health, Ohio State University, Columbus, Ohio.

PURPOSE. The retinal homeobox (Rx) gene plays an essential role in retinal development. An Rx-like (Rx-L) gene from Xenopus laevis has been identified. The purpose of this study was to analyze the function of Rx-L in the developing retina.

METHODS. DNA-binding properties of Rx-L were analyzed by electrophoretic mobility shift assay (EMSA), with in vitro–translated proteins and radiolabeled oligonucleotide probe. The Rx-L expression pattern was analyzed by in situ hybridization using whole or sectioned embryos and digoxigenin-labeled antisense riboprobes. Rx-L loss of function was studied by using antisense morpholino oligonucleotides targeted to the Rx-L translation initiation site. Embryos injected with control or Rx-L morpholinos were analyzed at stage 41 or 45.

RESULTS. Rx-L shares homology with Rx at the homeo-, OAR, and Rx domains, but lacks an octapeptide motif. Rx-L is expressed in the developing retina beginning in the early tailbud stage. In the maturing retina, Rx-L expression is restricted primarily to the developing photoreceptor layer and the ciliary marginal zone. Rx-L can bind a photoreceptor conserved element-1 (PCE-1) oligonucleotide, an element conserved among all known photoreceptor gene promoters. In a promoter activity assay, Rx-L functions as a stronger transcriptional activator than Rx. Antisense morpholino-mediated knockdown of Rx-L expression resulted in a decrease in rhodopsin and red cone opsin expression levels in Xenopus retinas. Injection of the Rx-L antisense morpholino oligonucleotide also resulted in a decrease in the length of both rod and cone outer segments.

CONCLUSIONS. The results suggest that Rx-L functions to regulate rod and cone development by activating photoreceptor-specific gene expression.






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Copyright © 2006 by the Association for Research in Vision and Ophthalmology