PUBLICATION

Mutation in the Zebrafish cct2 Gene Leads to Abnormalities of Cell Cycle and Cell Death in the Retina: A Model of CCT2-Related Leber Congenital Amaurosis

Authors
Minegishi, Y., Nakaya, N., Tomarev, S.I.
ID
ZDB-PUB-180223-27
Date
2018
Source
Investigative ophthalmology & visual science   59: 995-1004 (Journal)
Registered Authors
Keywords
none
MeSH Terms
  • Disease Models, Animal
  • Retina/pathology*
  • Chaperonin Containing TCP-1/genetics*
  • Amino Acid Sequence
  • Blotting, Western
  • Microscopy
  • Cell Cycle/genetics*
  • Apoptosis/genetics*
  • RNA/genetics
  • Zebrafish Proteins/genetics*
  • Zebrafish
  • In Situ Nick-End Labeling
  • Phenotype
  • Sequence Deletion*
  • CRISPR-Cas Systems
  • Molecular Sequence Data
  • Immunohistochemistry
  • Leber Congenital Amaurosis/genetics*
  • Animals
(all 19)
PubMed
29450543 Full text @ Invest. Ophthalmol. Vis. Sci.
Abstract
Purpose The compound heterozygous mutations in the ? subunit of chaperonin containing TCP-1 (CCT), encoded by CCT2, lead to the Leber congenital amaurosis (LCA). In this study, a cct2 mutant line of zebrafish was established to investigate the role of CCT2 mutations in LCA in vertebrates.
Methods A cct2 mutant zebrafish line was produced using the CRISPR-Cas9 system. Changes in the eyes of developing wild-type and mutant larvae were monitored using microscopy, immunostaining, TUNEL, and EdU assays. Phenotypic rescue of mutant phenotype was investigated by injection of CCT2 RNA into zebrafish embryos.
Results The cct2 mutation (L394H-7del) led to the synthesis of a mutated cct? protein with the L394H replacement and deletion of 7 amino acid residues (positions 395-401). The homozygous cct2-L394H-7del mutant exhibited a small eye phenotype at 2 days post fertilization (dpf) and was embryonically lethal after 5 dpf. In homozygous cct2-L394H-7del mutants, the retinal ganglion cell differentiation was attenuated, retinal cell cycle was affected, and the neural retinal cell death was significantly increased at 2 dpf compared with wild-type. Injection of RNA encoding wild-type human CCT? rescued the small eye phenotype, reduced retinal cell death, and restored the levels of CCT? protein and the major client protein G?1 that were significantly reduced in the homozygous cct2-L394H-7del mutant compared with wild-type. These results indicate that cct2 plays an essential role in retinal development by regulating the cell cycle.
Conclusions The retinal pathology observed in the homozygous cct2-L394H-7del mutants resembles the retinal pathology of human LCA patients.
Genes / Markers
Figures
Figure Gallery (3 images)
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Expression
Phenotype
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
zf3018
    		Indel
    1 - 1 of 1
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    Human Disease / Model
    Human Disease Fish Conditions Evidence
    Leber congenital amaurosisTAS
    1 - 1 of 1
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    Sequence Targeting Reagents
    Fish
    Fish
    cct2zf3018/zf3018
    WT
    1 - 2 of 2
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    Antibodies
    Orthology
    Engineered Foreign Genes
    No data available
    Mapping
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    Citation
    Minegishi, Y., Nakaya, N., Tomarev, S.I. (2018) Mutation in the Zebrafish cct2 Gene Leads to Abnormalities of Cell Cycle and Cell Death in the Retina: A Model of CCT2-Related Leber Congenital Amaurosis. Investigative ophthalmology & visual science. 59:995-1004.