Format

Send to

Choose Destination
Dev Biol. 2015 Dec 15;408(2):269-91. doi: 10.1016/j.ydbio.2015.03.010. Epub 2015 Mar 27.

An in vivo screen to identify candidate neurogenic genes in the developing Xenopus visual system.

Author information

1
Drug Discovery & Biomedical Sciences, The Medical University of South Carolina, Charleston, SC 29425, United States.
2
The Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA 92037, United States.
3
University of Nebraska Medical Center, Omaha, NE 68198, United States.
4
Dart Neuroscience, LLC, San Diego, CA 92064, United States.
5
The Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA 92037, United States. Electronic address: cline@scripps.edu.

Abstract

Neurogenesis in the brain of Xenopus laevis continues throughout larval stages of development. We developed a 2-tier screen to identify candidate genes controlling neurogenesis in Xenopus optic tectum in vivo. First, microarray and NanoString analyses were used to identify candidate genes that were differentially expressed in Sox2-expressing neural progenitor cells or their neuronal progeny. Then an in vivo, time-lapse imaging-based screen was used to test whether morpholinos against 34 candidate genes altered neural progenitor cell proliferation or neuronal differentiation over 3 days in the optic tectum of intact Xenopus tadpoles. We co-electroporated antisense morpholino oligonucleotides against each of the candidate genes with a plasmid that drives GFP expression in Sox2-expressing neural progenitor cells and quantified the effects of morpholinos on neurogenesis. Of the 34 morpholinos tested, 24 altered neural progenitor cell proliferation or neuronal differentiation. The candidates which were tagged as differentially expressed and validated by the in vivo imaging screen include: actn1, arl9, eif3a, elk4, ephb1, fmr1-a, fxr1-1, fbxw7, fgf2, gstp1, hat1, hspa5, lsm6, mecp2, mmp9, and prkaca. Several of these candidates, including fgf2 and elk4, have known or proposed neurogenic functions, thereby validating our strategy to identify candidates. Genes with no previously demonstrated neurogenic functions, gstp1, hspa5 and lsm6, were identified from the morpholino experiments, suggesting that our screen successfully revealed unknown candidates. Genes that are associated with human disease, such as such as mecp2 and fmr1-a, were identified by our screen, providing the groundwork for using Xenopus as an experimental system to probe conserved disease mechanisms. Together the data identify candidate neurogenic regulatory genes and demonstrate that Xenopus is an effective experimental animal to identify and characterize genes that regulate neural progenitor cell proliferation and differentiation in vivo.

KEYWORDS:

Candidate gene; Differentiation; In vivo imaging; Microarray; Morpholino; Neural progenitor cell; Neurogenesis; Proliferation; Radial glia

PMID:
25818835
PMCID:
PMC4584193
DOI:
10.1016/j.ydbio.2015.03.010
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center