Using the Tg(nrd:egfp)/albino zebrafish line to characterize in vivo expression of neurod.

Jennifer L Thomas,Peter F Hitchcock,Peter F Hitchcock,Margaret J Ochocinska,Margaret J Ochocinska,Ryan Thummel,Ryan Thummel

doi:10.1371/journal.pone.0029128

Jennifer L Thomas, Peter F Hitchcock + Show 5 more

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https://doi.org/10.1371/journal.pone.0029128

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Abstract

In this study, we used a newly-created transgenic zebrafish, Tg(nrd:egfp)/albino, to further characterize the expression of neurod in the developing and adult retina and to determine neurod expression during adult photoreceptor regeneration. We also provide observations regarding the expression of neurod in a variety of other tissues. In this line, EGFP is found in cells of the developing and adult retina, pineal gland, cerebellum, olfactory bulbs, midbrain, hindbrain, neural tube, lateral line, inner ear, pancreas, gut, and fin. Using immunohistochemistry and in situ hybridization, we compare the expression of the nrd:egfp transgene to that of endogenous neurod and to known retinal cell types. Consistent with previous data based on in situ hybridizations, we show that during retinal development, the nrd:egfp transgene is not expressed in proliferating retinal neuroepithelium, and is expressed in a subset of retinal neurons. In contrast to previous studies, nrd:egfp is gradually re-expressed in all rod photoreceptors. During photoreceptor regeneration in adult zebrafish, in situ hybridization reveals that neurod is not expressed in Müller glial-derived neuronal progenitors, but is expressed in photoreceptor progenitors as they migrate to the outer nuclear layer and differentiate into new rod photoreceptors. During photoreceptor regeneration, expression of the nrd:egfp matches that of neurod. We conclude that Tg(nrd:egfp)/albino is a good representation of endogenous neurod expression, is a useful tool to visualize neurod expression in a variety of tissues and will aid investigating the fundamental processes that govern photoreceptor regeneration in adults.

Highlights

NeuroD is a basic helix-loop-helix transcription factor that plays a common role in persistently mitotic cells as an essential link between cell cycle exit, cell fate determination, and cell survival [1]
The expression of the nrd:egfp transgene was first examined by wholemount fluorescence microscopy
We examined expression of the nrd:egfp transgene in relationship to retinal progenitors immunolabled with Proliferating Cell Nuclear Antigen (PCNA), a marker for proliferating cells [25,40]

Summary

Introduction

NeuroD is a basic helix-loop-helix (bHLH) transcription factor that plays a common role in persistently mitotic cells as an essential link between cell cycle exit, cell fate determination, and cell survival [1]. Neurod is expressed in the developing endocrine pancreas [5], the auditory and vestibular neuroblasts of the developing inner ear [6], and the lateral line of teleost fish [7] In both mice and zebrafish, neurogenin is expressed in cells prior to neurod, [2,4] and overexpression of the neurogenin homolog in Xenopus (X-NGNR-1) induces ectopic expression of Xneurod mRNA [8], suggesting that neurogenin is an upstream regulator of neurod. During both zebrafish and mammalian retinogenesis, neurod is first expressed in retinal neuroepithelial cells as they exit the cell cycle. Neurod expression was previously reported to persist in a subset of amacrine cells nascent cone photoreceptors near the retinal margins [1,10]

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