Loss of Mgat5a-mediated N-glycosylation stimulates regeneration in zebrafish
Wuhong Pei1, Sunny C Huang1, Lisha Xu1
1Functional and Translation Genome Branch, National Human Genome Research Institute, 9000 Rockville Pike, Building 50, Room 5537, Bethesda, MD 20892 USA.
Background:
We are using genetics to identify genes specifically involved in hearing regeneration. In a large-scale genetic screening, we identified mgat5a, a gene in the N-glycosylation biosynthesis pathway whose activity negatively impacts hair cell regeneration.
Methods:
We used a combination of mutant analysis in zebrafish and a hair cell regeneration assay to phenotype the loss of Mgat5a activity in zebrafish. We used pharmacological inhibition of N-glycosylation by swansonine. We also used over-expression analysis by mRNA injections to demonstrate how changes in N-glycosylation can alter cell signaling.
Results:
We found that mgat5a was expressed in multiple tissues during zebrafish embryo development, particularly enriched in neural tissues including the brain, retina, and lateral line neuromasts. An mgat5a insertional mutation and a CRISPR/Cas9-generated truncation mutation both caused an enhancement of hair cell regeneration which could be phenocopied by pharmacological inhibition with swansonine. In addition to hair cell regeneration, inhibition of the N-glycosylation pathway also enhanced the regeneration of lateral line axon and caudal fins. Further analysis showed that N-glycosylation altered the responsiveness of TGF-beta signaling.
Conclusions:
The findings from this study provide experimental evidence for the involvement of N-glycosylation in tissue regeneration and cell signaling.
