Effects of Homeobox Genes on the Differentiation of Photoreceptor and Nonphotoreceptor Neurons

• Published in: Investigative ophthalmology & visual science Vol. 43; no. 11; pp. 3522 - 3529
• Main Authors: Toy, Jeffrey, Norton, James S, Jibodh, Stefan R, Adler, Ruben
• Format: Journal Article
• Language: English
• Published: Rockville, MD ARVO 01-11-2002; Association for Research in Vision and Ophtalmology
• Subjects: Animals; beta-Galactosidase - genetics; Biological and medical sciences; Cell Culture Techniques; Cell Differentiation - physiology; Chick Embryo; Eye and associated structures. Visual pathways and centers. Vision; Eye Proteins - physiology; Fluorescent Antibody Technique, Indirect; Fundamental and applied biological sciences. Psychology; Green Fluorescent Proteins; Homeodomain Proteins - physiology; Luminescent Proteins - genetics; Neurons - cytology; Neurons - metabolism; Paired Box Transcription Factors; PAX6 Transcription Factor; Photoreceptor Cells, Vertebrate - cytology; Photoreceptor Cells, Vertebrate - metabolism; Repressor Proteins; Trans-Activators - physiology; Transcription Factors - physiology; Transfection; Vertebrates: nervous system and sense organs; Cell culture; Embryo; Photoreceptor; Cell differentiation; Visual system; Vertebrata; Neuron; Gene; Animal; Physiology; Chicken; Aves; Homeobox sequence; Progenitor cell
• ISSN: 0146-0404; 1552-5783

The homeobox genes Pax6 and Chx10 are diffusely expressed in proliferating, undifferentiated retina neuroepithelial cells. Distinct, topographically specific expression patterns emerge, however, as postmitotic cells become organized into layers. The hypothesis that the product of each gene may be necessary for the differentiation of particular nonphotoreceptor neuron subsets and that their absence may be required for progenitor cells to differentiate as photoreceptors was tested in this study.
Neural retinas from 5-day-old chick embryos were dissociated, cultured at low density, and cotransfected with a plasmid expressing the green fluorescent protein (GFP) reporter gene, and a plasmid expressing Pax6, Chx10, Optx2, or the control gene lacZ. After further culture, the cells were fixed and processed for the detection of cell-specific markers.
Nonphotoreceptor neurons increased threefold with Chx10 and almost sixfold with Pax6, compared with cells transfected with lacZ. The frequency of GFP(+) cells immunoreactive with the ganglion cell-specific antibody RA4 was unchanged by Chx10, but was increased twofold by Pax6. Conversely, Chx10 and Pax6 expression diminished the photoreceptor population to approximately 35% and 15% of control values, as determined by morphologic analysis, visinin immunocytochemistry, and peanut lectin binding. Optx2 had some inhibitory effects on photoreceptor differentiation, which were accompanied by marked increases in the frequency of morphologically undifferentiated cells.
These results are consistent with the hypothesis that Chx10 and Pax6 promote the differentiation of nonphotoreceptor neurons while inhibiting the differentiation of photoreceptor cells.