Figure 2

Figure 2

Zebrafish loss-of-function model for human SYNGAP1-RD. (A) Mammalian SYNGAP1 protein (H. sapiens and M. musculus) has four main protein interacting domains; pleckstrin homology (PH) domain, C2 domain, RasGAP domain, and coiled-coiled (CC) domain. Zebrafish (D. rerio) Syngap1 orthologs; Syngap1a and Syngap1b show domain conservation with that of mammals. (B) Syngap1ab protein diagrams show sites of CRISPR induced mutations. Resulting CRISPR mutants used for phenotypic analyses: syngap1a¹ allele p.Ser43Argfs*21 and syngap1b allele p.Met149Ilefs*9. syngap1a¹ had an amino acid change from a serine to an arginine at position 43 introducing a premature stop codon, 21 amino acids downstream. Syngap1b mutant allele had a change of methionine to an isoleucine at position 49 introducing a premature stop codon 9 amino acids downstream. (C) Western blots illustrate the expression of SYNGAP1 in whole brain lysates from adult mice and zebrafish. In wild-type zebrafish, the Syngap1 protein was detected at a similar molecular weight (~150 kDa) to that of the mouse SYNGAP1, using a rat anti-Syngap1 antibody. GAPDH and tubulin were used as the loading controls for mouse and zebrafish, respectively. (D) Mutant syngap1ab larvae showed reduced syngap1a and syngap1b mRNA expression levels at 7dpf. Group comparisons were made using 2-way ANOVA followed by Tukey’s multiple comparison test. p value asterisks represent: *p < 0.05, **p < 0.01; ***p < 0.001; ****p < 0.0001.