Construction and otological phenotypic characterization of a mouse model harboring orthologous human Waardenburg syndrome mutation SOX10 p.R106W

Objective To compare the otological phenotypic characteristics induced by Waardenburg syndrome mutation SOX10 p.R106W in mouse, miniature pig and human, and to construct a mouse model harboring orthologous human SOX10 p.R106W mutation. Methods By comparing the amino acid sequences of SOX10 proteins from human, miniature pig and mouse, the mouse orthologous site corresponding to human SOX10 p.R106W mutation was identified, and the nucleic acid mutation Sox10 c.316A > T coding Sox10 p.R106W was precisely introduced into the orthologous site of mouse Sox10 coding sequence by CRISPR/Cas9-induced homologous recombination. Characteristics of otological phenotypes were assessed by auditory brainstem response test and cochlear tissue histology. mRNA sequencing was conducted on cochlear tissues, and differential expressed genes (DEGs) were accepted gene ontology (GO) analysis. The correlation between DEGs and Sox10 was analyzed through String database. Results The gene-edited mice harboring Sox10 c.316A > T (Sox10 p.R106W) mutation were successfully generated. The skin of gene-edited mice exhibited white spots and stable heterozygous inheritance. Sox10 c.316A > T/+ mutant mice showed no notable difference in cochlea histological structure as well as response to acoustic stimulation, compared with wild type (WT) littermates. Transcriptomic analysis revealed that Sox10 c.316A > T/+ mutant mice had 323 upregulated genes in concomitant with 283 downregulated genes. GO analysis showed that upregulated genes were associated with immunity while downregulated genes were associated with neurological functions. Five genes, neurofilament heavy polypeptide(Nefh)、fatty acid 2-hydroxylase(Fa2h)、gap junction protein beta 1(Gjb1)、nerve growth factor receptor(Ngfr) and zinc finger protein 536(Zfp536) were correlated with Sox10, but had no relationship with cochlear development and function. Conclusion Sox10 p.R106W mutant mouse model is successfully established using CRISPR/Cas9 system. Compared with WT mice, the gene-edited mice have no difference in otological phenotypes, while there is significant difference in disease phenotype induced by these mutation in human and miniature pig.