Exome Sequencing and Functional Validation in Zebrafish Identify GTDC2 Mutations as a Cause of Walker-Warburg Syndrome

Abstract

Whole-exome sequencing (WES), which analyzes the coding sequence of most annotated genes in the human genome, is an ideal approach to studying fully penetrant autosomal-recessive diseases, and it has been very powerful in identifying disease-causing mutations even when enrollment of affected individuals is limited by reduced survival. In this study, we combined WES with homozygosity analysis of consanguineous pedigrees, which are informative even when a single affected individual is available, to identify genetic mutations responsible for Walker-Warburg syndrome (WWS), a genetically heterogeneous autosomal-recessive disorder that severely affects the development of the brain, eyes, and muscle. Mutations in seven genes are known to cause WWS and explain 50%-60% of cases, but multiple additional genes are expected to be mutated because unexplained cases show suggestive linkage to diverse loci. Using WES in consanguineous WWS-affected families, we found multiple deleterious mutations in GTDC2 (also known as AGO61). GTDC2's predicted role as an uncharacterized glycosyltransferase is consistent with the function of other genes that are known to be mutated in WWS and that are involved in the glycosylation of the transmembrane receptor dystroglycan. Therefore, to explore the role of GTDC2 loss of function during development, we used morpholino-mediated knockdown of its zebrafish ortholog, gtdc2. We found that gtdc2 knockdown in zebrafish replicates all WWS features (hydrocephalus, ocular defects, and muscular dystrophy), strongly suggesting that GTDC2 mutations cause WWS.

Genetics and mechanisms leading to human cortical malformations

“…Mutant mice of the integrin-related family, such as FAK (focal adhesion kinase) [233] and Ilk (integrin-linked kinase) [235], have been associated with features that resemble human COB-LIS, including cortical migration and basal lamina integrity defects, but mutations of these genes have not been identified in human patients.…”

Gènes impliqués dans les alpha-dystroglycanopathies

“…Il est également impliqué dans des cas de LISII foetale [2]. Le gène GTDC2 est, pour sa part, impliqué dans des formes de LISII sévères, dans des cas de WWS post-natal ou des formes moins sévères de LGMD [19]. Enfin, un cinquième gène, B3GALNT2, code une glycosyltransférase du réticulum endoplasmique.…”

The O-Mannosylation Pathway: Glycosyltransferases and Proteins Implicated in Congenital Muscular Dystrophy

“…Defects in this gene are seen in patients presenting with Walker-Warburg syndrome, and knockdown of this gene in zebrafish recapitulated the phenotype of the knockdown of POMT1 (75). It was speculated that this enzyme might be a xylosyltransferase (GT61 does contain ␤1,2-xylosyltransferases) (75). Sequence comparison showed that this enzyme also has high homology to the recently described secretory pathway-localized protein O-␤-N-acetylglucosaminyltransferase (EGF domain-specific O-GlcNAc transferase) (76,77).…”

“…This glycosyltransferase belongs to the CAZy inverting GT61 family. Defects in this gene are seen in patients presenting with Walker-Warburg syndrome, and knockdown of this gene in zebrafish recapitulated the phenotype of the knockdown of POMT1 (75). It was speculated that this enzyme might be a xylosyltransferase (GT61 does contain ␤1,2-xylosyltransferases) (75).…”

“…Glycosyltransferase-like Domain-containing 2 Protein (GTDC2)-The most recent protein associated with CMD is GTDC2 (75). This glycosyltransferase belongs to the CAZy inverting GT61 family.…”