Abstract
Purpose:
The transcription factor neural retina leucine zipper (NRL) is essential to specify rod photoreceptor cell fate during retina development, as targeted deletion of Nrl (-/-) in mice led to a complete absence of rod photoreceptors. Nrl -/- rod precursors fail to turn on rod genes and differentiate into functional cone photoreceptors. Previous ChIP-seq analysis of mouse retina revealed NRL binding to the intron of the receptor accessory protein 6 (Reep6). Our goal is to elucidate the mechanism of transcriptional regulation of the Reep6 gene and to understand the molecular mechanism of NRL action.
Methods:
We performed morpholino experiments in zebra fish to test the functional relevance of Reep6 in retina development. To study the transcriptional regulation of Reep6 gene, we used a combined approach including rapid analysis of cDNA ends (5’-RACE), chromatin immunoprecipitation, luciferase reporter assay, promoter mapping in retina explant and in vivo shRNA analysis.
Results:
Knockdown of Reep6 in zebrafish by morpholinos led to microphthalmia. NRL binding to the Reep6 intron enhanced its promoter activity in a luciferase reporter assay and also in a GFP reporter assay in retina explant.
Conclusions:
Reep6 expression is controlled by NRL through an enhancer located within its intron. NRL direct target genes serve as excellent candidates for retinal disease gene discovery.
Keywords: 738 transcription •
539 genetics •
648 photoreceptors