Dystroglycanopathies (DGs) are a group of rare autosomal recessive congenital muscular dystrophies (CMDs) that present at birth and severely affect the brain, eyes, and muscles. Therapies for DGs have been stymied by their high genetic and clinical heterogeneity. Furthermore, mouse knock-out (KO) models have only proven partially suitable for preclinical studies due to embryonic lethality and must be generated by conditional tissue approaches. As such, there is a critical need for a complete KO which can recapitulate the human disease throughout development. To overcome these limitations, we characterized an alternative model. Zebrafish have genetic conservation and are an attractive vertebrate for high throughput screenings.
Most DGs genes converge on defects of glycosylation of the glycoprotein alpha-dystroglycan (alpha-DG). Without alpha-DG glycosylation, transmembrane communication between the cell and extracellular matrix (ECM) is compromised. Most DG genes are glycosyltransferases participating in the assembly of a glycan chain on alpha-DG which binds ECM components in multiple tissues. The initial step in glycosylation is the addition of an O-linked mannose via (POMT1) to alpha-DG. POMT1 mutations in patients are frequently the most severe form of CMD with early lethality, Walker-Warburg Syndrome. Our zebrafish strain has a premature stop codon leading to complete loss of POMT1 and alpha-DG glycosylation. Investigation showed that loss of pomt1 leads to early lethality starting at 30 days post fertilization (dpf), small size, impaired mobility, muscle disease and retinal defects. However, these phenotypes are delayed compared to dystroglycan (dag1) KO in fish. Since pomt1 is critical for early development and mRNA is provided by the mother in the yolk, we asked whether KO embryos obtained from KO mothers would have more severe phenotypes. Zebrafish larvae from KO X heterozygote crosses begin to die at 10 dpf similar to dag1 KO zebrafish and have mobility issues at 5 dpf. Furthermore, muscle disease, retinal synapse formation deficits, and axon guidance defects were uncovered during the first week by generating KO embryos from KO mothers. Our findings show that when the contribution of maternal mRNA is properly defined, DGs mutations in the zebrafish lead to disease phenotypes consistent with the human presentation. With this high-throughput model system, our research can both unravel and overcome genetic-phenotypic heterogeneity for treatment.