Neural crest cells regulate optic cup morphogenesis by promoting extracellular matrix assembly

Chase Dallas Bryan, Rebecca Lynne Pfeiffer, Bryan William Jones, Kristen Marie Kwan

Preprint posted on July 22, 2018

Ocular Origami: Bryan and colleagues discover that in the developing zebrafish eye, cell and tissue movements in the optic cup are dependent on the extracellular matrix protein nidogen provided by surrounding neural crest cells.

Selected by Ashrifia Adomako-Ankomah

During embryogenesis, eye development is marked by several critical cell and tissue movement events. As reviewed by Cavodeassi (2018), zebrafish eye development begins with the specification of the eye field cells. These cells then evaginate to form the optic vesicles. The distal edges of the optic vesicles subsequently invaginate, and the optic cup is formed. The optic cup has two layers- the outer layer forms the retinal pigment epithelium (RPE), and the inner layer forms the neural retina. The developing optic cup is surrounded by a periocular mesenchyme (POM) made up of neural crest and mesoderm-derived mesenchyme cells. The POM is known to influence cell migration during eye development, though the exact mechanisms by which this happens are not fully resolved. In this preprint, Bryan and colleagues take advantage of advances in microscopy and live imaging technology to examine in detail the role of the POM neural crest cells in regulating tissue movements during optic cup morphogenesis.

Interesting findings
The authors first set out to determine the location of neural crest cells in the region of the developing eye. For this, they employ the vast transgenic toolkit available to zebrafish researchers by crossing two fish lines: one in which GFP labels all cell membranes, and the other in which RFP labels only neural crest membranes. Results from 4-dimensional time lapse imaging in these double-labeled fish show the gradual migration of neural crest cells around and in close contact with the developing optic cup until they surround the RPE.


Preprint Supplemental Movie 1

To identify the function of the neural crest cells, eye development was observed in two mutant fish lines (the tfap2a;foxd3 double mutant and the paf1 mutant) in which these cells were depleted. When compared to wild type siblings, neural crest mutants had defects in early optic cup invagination, leading to malformations of the eye.

                 Preprint Figure 2

The authors then used confocal imaging to track the migration paths of individual cells to figure out exactly which cell movements were regulated by neural crest cells. Results showed that migrating cells in mutant embryos did not follow the stereotypical trajectory seen in wild type embryos. In neural crest mutants, these cells traveled farther, faster, and were found in a wider area than cells in control embryos.

How do neural crest cells control cell migration? As the basement membrane of the RPE was disorganized in neural crest mutants, it was plausible that some component of the ECM was missing in the absence of neural crest cells. In situ hybridization data revealed that nidogen, an ECM crosslinking protein known to regulate basement membrane assembly, was expressed in neural crest cells surrounding the RPE and lost in neural crest mutants. Blocking the function of nidogen perturbed lens morphogenesis, and the general overexpression of nidogen partially rescued eye defects in neural crest mutants. Additionally, the overexpression of nidogen in a wild type background also led to optic cup invagination defects.

The authors conclude that nidogen produced by the neural crest cells is required for the proper formation of the basement membrane of the RPE. This creates a restrictive environment, helping cells in the optic vesicle migrate to the right location as the optic cup invaginates.

      Preprint Figure 10

What I like about this preprint
This preprint makes good use of the ability to make stable mutant and transgenic lines in zebrafish. The gorgeous confocal images shown provide great detail on a single-cell level, and the movies are fascinating to watch. Also, imaging data were captured prior to genotyping the embryos used, ensuring objectivity in data collection.

Follow-up question
What do the authors speculate is the role, if any, of the mesoderm-derived mesenchyme cells of the POM in regulating optic cup morphogenesis?

1. Cavodeassi, F. (2018) Dynamic Tissue Rearrangements during Vertebrate Eye Morphogenesis: Insights from Fish Models. Journal of Developmental Biology 6(1), 4

Tags: cell migration, extracellular matrix, eye cup morphogenesis, eye development, neural crest cells, nidogen, zebrafish

Posted on: 30th August 2018

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  • Author's response

    Dr. Kristen M. Kwan shared

    We agree that the follow-up question you’ve raised is indeed a very interesting one (and one we are interested in pursuing in the future).

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