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Published on: September 1, 2017
Zebrafish cdc6 hypomorphic mutation causes Meier-Gorlin syndrome-like phenotype
Likun Yao1, Jing Chen1, Xiaotong Wu1
1Laboratory of Molecular Developmental Biology, State Key Laboratory of Membrane Biology, Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Abstract:
Cell Division Cycle 6 (Cdc6) is a component of pre-replicative complex (preRC) forming on DNA replication origins in eukaryotes. Recessive mutations in ORC1, ORC4, ORC6, CDT1 or CDC6 of the preRC in human cause Meier-Gorlin syndrome (MGS) that is characterized by impaired post-natal growth, short stature and microcephaly. However, vertebrate models of MGS have not been reported. Through N-ethyl-N-nitrosourea mutagenesis and Cas9 knockout, we generate several cdc6 mutant lines in zebrafish. Loss-of-function mutations of cdc6, as manifested by cdc6tsu4305 and cdc6tsu7cd mutants, lead to embryonic lethality due to cell cycle arrest at the S phase and extensive apoptosis. Embryos homozygous for a cdc6 hypomorphic mutation, cdc6tsu21cd, develop normally during embryogenesis. Later on, compared with their wild-type (WT) siblings, cdc6tsu21cd mutant fish show growth retardation, and their body weight and length in adulthood are greatly reduced, which resemble human MGS. Surprisingly, cdc6tsu21cd mutant fish become males with a short life and fail to mate with WT females, suggesting defective reproduction. Overexpression of Cdc6 mutant forms, which mimic human CDC6(T323R) mutation found in a MGS patient, in zebrafish cdc6tsu4305 mutant embryos partially represses cell death phenotype, suggesting that the human CDC6(T323R) mutation is a hypomorph. cdc6tsu21cd mutant fish will be useful to detect more tissue defects and develop medical treatment strategies for MGS patients.
Insights
Zebrafish cdc6 mutants reveal new insights into Meier-Gorlin syndrome (MGS). A hypomorphic mutation causes growth defects and reproductive issues, mimicking human MGS and offering a valuable vertebrate model for research.
Area of Science:
- Genetics
- Developmental Biology
- Molecular Biology
Background:
- Meier-Gorlin syndrome (MGS) is a human genetic disorder linked to mutations in pre-replicative complex (preRC) genes, including CDC6.
- Vertebrate models for MGS are currently lacking, hindering research into the syndrome's mechanisms and potential treatments.
Purpose of the Study:
- To generate and characterize zebrafish models of MGS by creating cdc6 mutant lines.
- To investigate the in vivo function of Cdc6 in vertebrate development and its role in MGS pathogenesis.
Main Methods:
- Zebrafish were subjected to N-ethyl-N-nitrosourea mutagenesis and Cas9 knockout to generate cdc6 mutant lines.
- Phenotypic analysis of loss-of-function and hypomorphic cdc6 mutants was performed, including assessment of embryonic lethality, cell cycle progression, apoptosis, growth, and reproductive capacity.
- Human CDC6 mutations were modeled by overexpression in zebrafish mutants to assess functional complementation.
Main Results:
- Loss-of-function cdc6 mutants exhibited embryonic lethality due to S-phase arrest and apoptosis.
- A hypomorphic cdc6 mutant (cdc6tsu21cd) displayed growth retardation, reduced adult size, and reproductive defects, closely resembling human MGS.
- Overexpression of a human CDC6 mutation (CDC6(T323R)) partially rescued the cell death phenotype in zebrafish mutants, suggesting it is a hypomorphic mutation.
Conclusions:
- Zebrafish cdc6 mutants provide a valuable vertebrate model for studying MGS.
- The cdc6tsu21cd mutant fish exhibit phenotypes relevant to MGS, including growth and reproductive abnormalities.
- This model can facilitate further investigation into MGS tissue defects and the development of therapeutic strategies.
