Inheritance of the epigenetic signature and reduced intermuscular bone phenotype acquired via DNA methylation editing of the runx2b promoter in zebrafish
- 1Institute of Modern Aquaculture Science and Engineering, Guangdong-Macao Joint Laboratory for Aquaculture Breeding Development and Innovation, School of Life Sciences, South China Normal University, Guangzhou, Guangdong 510631, China
- 2Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology, School of Life Sciences, South China Normal University, Guangzhou, Guangdong 510631, China
- 3Fisheries College, Ocean University of China, Qingdao, Shandong 266003, China
Abstract
The presence of intermuscular bones (IBs) can directly affect the economic value of aquaculture fish. Although genome editing can create IB-free fish by knocking out key IB-related genes, such as runx2b, the associated DNA sequence alterations raise food safety and health concerns, limiting its breeding applications. In this study, we used CRISPR/dCas9-mediated epigenome-editing technology targeting the runx2b promoter in zebrafish to alter DNA methylation patterns without changing the DNA sequence. Our results showed that higher runx2b promoter methylation patterns significantly inhibited eGFP mRNA expression levels in the recombinant plasmid. Using the CRISPR/dCas9-Dnmt7 system to enhance methylation of the zebrafish runx2b promoter, we observed a significant decrease in runx2b mRNA expression levels in the F0 generation. The IBs in the 11th–16th muscle segments of the adult F0 fish were significantly shorter compared with the controls. Inbreeding of fish was used to produce F1 and F2 offspring that retained these high promoter methylation levels, along with persistent runx2b expression suppression and IB development inhibition. Transcriptome sequencing analysis suggested that increasing runx2b promoter methylation levels may synergistically induce additional epigenetic modifications, potentially affecting the PPAR signaling pathway and FoxO transcription factor regulation, which appears to inhibit osteoblast proliferation and differentiation. Overall, this study demonstrates an innovative application of epigenetic editing technology for aquaculture breeding. By precisely regulating the expression patterns of key genes for economically important traits while preserving genomic DNA integrity, this approach provides a theoretical foundation and technical support for improving fish economic traits.
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BibTeX
@article{liang2026inheritance,
title = {{Inheritance of the epigenetic signature and reduced intermuscular bone phenotype acquired via DNA methylation editing of the runx2b promoter in zebrafish}},
author = {Fang Liang and Bi-Yu Wu and Jia-Yi Liu and Xue Wang and Min-Xing Liang and Chao-Yi Wang and Jing-Yi Zhang and Jun-Jie Wang and Yu-Tao Miao and Kang-Sen Mai and Jun Zhao and Xue-Geng Wang},
journal = {Zoological Research},
year = {2026},
month = sep,
doi = {10.24272/j.issn.2095-8137.2025.494},
url = {https://doi.org/10.24272/j.issn.2095-8137.2025.494},
note = {CC BY-NC 4.0},
}
RIS
TY - JOUR
TI - Inheritance of the epigenetic signature and reduced intermuscular bone phenotype acquired via DNA methylation editing of the runx2b promoter in zebrafish
AU - Fang Liang
AU - Bi-Yu Wu
AU - Jia-Yi Liu
AU - Xue Wang
AU - Min-Xing Liang
AU - Chao-Yi Wang
AU - Jing-Yi Zhang
AU - Jun-Jie Wang
AU - Yu-Tao Miao
AU - Kang-Sen Mai
AU - Jun Zhao
AU - Xue-Geng Wang
PY - 2026
DA - 2026/09/18
JO - Zoological Research
DO - 10.24272/j.issn.2095-8137.2025.494
UR - https://doi.org/10.24272/j.issn.2095-8137.2025.494
AB - The presence of intermuscular bones (IBs) can directly affect the economic value of aquaculture fish. Although genome editing can create IB-free fish by knocking out key IB-related genes, such as runx2b, the associated DNA sequence alterations raise food safety and health concerns, limiting its breeding applications. In this study, we used CRISPR/dCas9-mediated epigenome-editing technology targeting the runx2b promoter in zebrafish to alter DNA methylation patterns without changing the DNA sequence. Our results showed that higher runx2b promoter methylation patterns significantly inhibited eGFP mRNA expression levels in the recombinant plasmid. Using the CRISPR/dCas9-Dnmt7 system to enhance methylation of the zebrafish runx2b promoter, we observed a significant decrease in runx2b mRNA expression levels in the F0 generation. The IBs in the 11th–16th muscle segments of the adult F0 fish were significantly shorter compared with the controls. Inbreeding of fish was used to produce F1 and F2 offspring that retained these high promoter methylation levels, along with persistent runx2b expression suppression and IB development inhibition. Transcriptome sequencing analysis suggested that increasing runx2b promoter methylation levels may synergistically induce additional epigenetic modifications, potentially affecting the PPAR signaling pathway and FoxO transcription factor regulation, which appears to inhibit osteoblast proliferation and differentiation. Overall, this study demonstrates an innovative application of epigenetic editing technology for aquaculture breeding. By precisely regulating the expression patterns of key genes for economically important traits while preserving genomic DNA integrity, this approach provides a theoretical foundation and technical support for improving fish economic traits.
SN - 2095-8137
ER -