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Scott Taylor, James Fadool, Ryan Thummel, Peter Hitchcock; bHLH transcription factor NeuroD is a non-cell autonomous regulator of cell cycle withdrawal during photoreceptor development and regeneration in the zebrafish retina. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1165.
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© ARVO (1962-2015); The Authors (2016-present)
Development and regeneration in the central nervous system require precise regulation of cell cycle entry and exit. In the retina of the embryonic zebrafish, the bHLH transcription factor NeuroD mediates exit of photoreceptor progenitors from the cell cycle (Ochocinska and Hitchcock, 2009). In adult zebrafish, NeuroD is expressed in photoreceptor progenitors during injury-induced regeneration (Thomas et al., 2012), but its function in this process was previously not known. The purpose of this research was to investigate NeuroD expression and function during photoreceptor regeneration in adult zebrafish.
First, co-expression studies were performed with genes expressed in retinal stem cells and cells of the photoreceptor lineage. Second, NeuroD loss-of-function and gain-of-function approaches were used to test the hypothesis that during the selective regeneration of photoreceptors NeuroD regulates cell cycle withdrawal in photoreceptor progenitors. Third, genetic mosaic analysis was performed to gain insights into the mechanisms by which NeuroD functions during photoreceptor genesis and regeneration.
Co-expression studies revealed that in the regenerating retina, neuroD is expressed in pax6-negative mitotic photoreceptor progenitors and nascent photoreceptors derived from Muller glia. These progenitors express markers of canonical Wnt and TGF-beta signaling pathways, and later express crx and nr2e3. NeuroD loss-of-function in regenerating retinas (morpholino-induced NeuroD knockdown) resulted in reduced rod cell regeneration by 7 days post-lesioning (dpl). Systemic labeling with EdU (5-ethynyl-2-deoxyuridine) at 3 dpl and BrdU (5-bromo-2-deoxyuridine) at 6 dpl revealed that by 6 dpl fewer progenitors exit the cell cycle in NeuroD knockdown retinas. Finally, the genetic chimeric analysis (NeuroD+ donor/NeuroD- recipient and NeuroD- donor/NeuroD+ recipient embryos) revealed that NeuroD function is mediated through non-cell autonomous mechanisms in the embryonic retina.
Together, these data show that during photoreceptor development and regeneration, NeuroD is required for photoreceptor progenitors to exit the cell cycle and differentiate. Importantly, the function of NeuroD in the developing retina is non-cell autonomous, suggesting that secreted factors mediate NeuroD function.
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