Knockout of fcsk gene in zebrafish causes neurodevelopmental defects

Congenital disorders of glycosylation (CDG) are a cluster of single-gene disorders caused by defects in glycosylation modification. FCSK encodes fucokinase, which converts L-fucose to fucose-1-phosphate and plays an important role in fucosylated modifications. The FCSK mutations cause CDG with autosomal recessive inheritance pattern, characterized primarily by developmental delay, hypotonia, and brain abnormalities. Currently, there are no fcsk mutant animal models. In this study, we constructed the first fcsk knockout (fcsk-/-) zebrafish by CRISPR/cas9 technology. fcsk-/- zebrafish exhibited stunted growth, characterized by delayed epiboly and DNA accumulation in early developmental stages, and brain atrophy in adult stage. Larvae fcsk-/- zebrafish exhibited impaired locomotion and increased the susceptibility to pentylenetetrazole-induced seizures. Adult fcsk-/- zebrafish showed neurodevelopmental disorders, including increased anxiety, decreased aggression, reduced social preference, and impaired memory. Additionally, the fucosylation level of total protein was reduced in fcsk-/- zebrafish. The pofut2 gene encodes protein O-fucosyltransferase 2, which is involved in the salvage pathway of fucosylation, and the expression of pofut2 was reduced in fcsk-/- zebrafish. Apoptosis in the midbrain-hindbrain boundary (MHB) cells was increased in fcsk-/- zebrafish. The developmental defects and total protein fucosylation level in fcsk-/- zebrafish can be rescued by supplementing with GDP-L-fucose or human FCSK gene. RNA-sequencing further indicated that disorders may be attributed to altered gene expression levels associated with glycosylation, apoptosis, and neurodegenerative diseases. In conclusion, our findings suggest that fcsk-/- zebrafish exhibit neurodevelopmental disorders, which provides first fcsk gene knockout animal model for future studies on molecular mechanisms and drug screening.