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A. deCarvalho, J.M. Fadool; Cloning and Expression Pattern of Pou Domain Transcription Factors in the Developing Zebrafish Visual System . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1661.
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Purpose: Murine members of the class IV POU domain transcription factors are expressed in the developing retina and superior colliculus. From mouse knockout studies, it has been shown that Brn-3b (Brn 3.2, Pou4f2) is essential for the normal differentiation and maturation of retinal ganglion cells (RGC). We cloned and characterized the spatio-temporal expression pattern of the zebrafish Brn-3b homologue as a first step in understanding its function in zebrafish retinal and tectal development. Methods: A combination of PCR-based DNA walking and rapid amplification of cDNA ends (RACE) was used to clone the complete coding region of the two isoforms of zebrafish Brn-3b. Brn-3b(l) and Brn-3b(s) transcripts were detected by RT-PCR from zebrafish embryos and adult tissues. Spatial distribution of Brn-3b and Brn-3c transcripts during development was assessed using whole mount in situ hybridization. Results: The deduced amino acid sequence of zebrafish Brn-3b shows 100% and 99% identity with mouse and human sequences for the POU IV box and POU domain, respectively. Both Brn-3b transcripts were detected by RT-PCR from early retinal development (36 hours post fertilization), through a stage where the zebrafish visual system is functional (5 days post-fertilization). Brn-3b transcripts were also detected in adult tectum and, at lower levels, in adult retina. In situ hybridization results showed expression of Brn-3b in RGC and tectum during development. In contrast, Brn-3c did not co-localize with the Brn-3b transcripts during development, but was expressed in adult retina and tectum. Conclusion: Our results are in agreement with the expression of Brn-3b in the developing and adult murine visual system. In zebrafish, both Brn-3b transcripts seem to have similar expression pattern in the developing visual system.
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