Establishment and characterization of Neu1-knockout zebrafish and its abnormal clinical phenotypes

Okada, K., Takase, R., Hamaoka, Y., Honda, A., Ikeda, A., Hokazono, Y., Maeda, Y., Hayasaka, O., Kotani, T., Komatsu, M., Shiozaki, K.
The Biochemical journal   477(15): 2841-2857 (Journal)
Registered Authors
Kotani, Tomoya, Shiozaki, Kazuhiro
Neu1 sialidase, genome editing, lysosome, zebrafish
MeSH Terms
  • Animals
  • Animals, Genetically Modified
  • Body Weight/genetics
  • CRISPR-Cas Systems
  • Disease Models, Animal
  • Embryo, Nonmammalian
  • Female
  • Gene Expression Regulation, Developmental
  • Gene Knockout Techniques
  • Glycoproteins/genetics
  • Glycoproteins/metabolism
  • HEK293 Cells
  • Humans
  • Hydrogen-Ion Concentration
  • Male
  • Mucolipidoses/etiology
  • Mucolipidoses/genetics
  • N-Acetylneuraminic Acid/metabolism
  • Neuraminidase/genetics*
  • Neuraminidase/metabolism*
  • Osteogenesis/genetics
  • Phenotype
  • Zebrafish/embryology
  • Zebrafish/genetics*
  • Zebrafish Proteins/genetics*
  • Zebrafish Proteins/metabolism
32686823 Full text @ Biochem. J.
Mammalian sialidase Neu1 is involved in various physiological functions, including cell adhesion, differentiation, cancer metastasis, and diabetes through lysosomal catabolism and desialylation of glycoproteins at the plasma membrane. Various animal models have been established to further explore the functions of vertebrate Neu1. The present study focused on zebrafish (Danio rerio) belonging to Cypriniformes as an experimental animal model with neu1 gene deficiency. The results revealed that the zebrafish Neu1 desialyzed both a2-3 and a2-6 sialic acid linkages from oligosaccharides and glycoproteins at pH 4.5, and it is highly conserved with other fish species and mammalian Neu1. Further, Neu1-knockout zebrafish (Neu1-KO) was established through CRISPR/Cas9 genome editing. Neu1-KO fish exhibited slight abnormal embryogenesis with the accumulation of pleural effusion; however, no embryonic lethality was observed. Although Neu1-KO fish were able to be maintained as homozygous, they showed smaller body length and weight than the wild type (WT) fish, and muscle atrophy and curvature of the vertebra were observed in adult Neu1-KO fish (8 months). The expression patterns of myod and myog transcription factors regulating muscle differentiation varied between Neu1-KO and WT fish embryo. Expression of lysosomal-related genes, including ctsa,lamp1a, and tfeb were upregulated in adult Neu1-KO muscle as compared to WT. Furthermore, the expression pattern of genes involved in bone remodeling (runx2a, runx2b, and mmp9) was decreased in Neu1-KO fish. These phenotypes were quite similar to those of Neu1-KO mice and human sialidosis patients, indicating the effectiveness of the established Neu1-KO zebrafish for the study of vertebrate Neu1 sialidase.
Genes / Markers
Show all Figures
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes