ZBED6 deficiency enhances porcine carcass traits and meat quality via ISLR-mediated Wnt signaling activation

doi:10.1016/j.jia.2025.04.018

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Huan Liu¹^{, 2*}, Jiahe Huang¹^*, Junying Xiao¹^*, Ruirui Pan¹, Ruixin Zhang¹, Xiangyang Xing³, Xiao Zhang¹, Bo Xia¹,Dengke Pan³^#, and Jiangwei Wu¹^#

¹Key Laboratory of Animal Genetics, Breeding and Reproduction of Shaanxi Province, College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi 712100, China.

²Department of Emergency, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.

³Chengdu Clonorgan Biotechnology Co.,ltd., Chengdu, 610000, China.

Highlights

l ZBED6 deficiency improves growth performance and meat quality of pigs.

l ZBED6 deficiency increases the content of n-3PUFA, EAA and FAA in LD muscle.

l ISLR is a ZBED6-IGF2-independent and evolutionarily conserved direct target of ZBED6.

l ZBED6 deficiency promotes skeletal muscle growth through activates the ISLR/Wnt pathway.

Abstract
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摘要

【目的】鉴定调控猪产肉性能与肉品质的关键基因对生猪产业可持续发展至关重要。本研究旨在系统解析转录因子锌指蛋白6（ZBED6）对猪胴体性状和肉品质的作用，并深入探究其分子调控机制，评估其缺失猪作为新型种质资源的应用潜力。【方法】基于ZBED6缺失猪模型，通过生长性能评估、肉品质测定和多组学分析等方法，活体水平解析ZBED6对骨骼肌生长的作用和机制。在C2C12成肌细胞中敲低Zbed6，离体水平验证ZBED6的靶基因和关键信号通路。最后，采用90天大鼠亚慢性毒性实验评估ZBED6缺失猪肉的安全性。【结果】与对照组相比，ZBED6缺失猪生长性能和胴体性能显著提高，其体重显著增加15.18%（P<0.01）、饲料转化率显著降低21.45%（P<0.01）、胴体瘦肉率提升6.21%（P<0.001）。肉品质和营养价值评估表明，ZBED6缺失猪的肉色红度值（a*）提高17.27%（P<0.05），蒸煮损失率降低2.00%（P<0.05），n-3多不饱和脂肪酸提高3.85%，必需氨基酸总量增加6.74%。联合RNA-seq、定量蛋白质组学和ChIP-seq等多组学方法，筛选出ISLR和Wnt/β-catenin信号通路是ZBED6的靶基因和下游信号通路；在C2C12细胞中敲低Zbed6，显著上调ISLR表达，并激活经典Wnt/β-catenin通路，促进成肌细胞分化，进一步验证ZBED6缺失通过负调控ISLR表达和激活Wnt/β-catenin通路促进骨骼肌生长并改善猪肉品质。最后，通过毒理实验表明长期饲喂ZBED6缺失猪肉对大鼠血液生化指标、器官系数等均无显著影响（P>0.05），证明ZBED6缺失猪肉的安全性。【结论】本研究首次证实ZBED6缺失通过促进ISLR的表达，激活Wnt信号通路从而促进生猪骨骼肌生长，系统阐明了其提升产肉效率和改善肉品质的分子机制。【创新性】本研究基于ZBED6缺失猪，首次揭示ZBED6缺失通过靶向抑制ISLR介导Wnt/β-catenin信号通路激活进而促进猪骨骼肌生长和改善肉品质的分子机制，实现生猪骨骼肌生长和猪肉品质的双向改良，为ZBED6基因编辑猪的未来生产应用提供了理论参考。

Abstract

Identification of genes that regulate meat production and quality in pigs is crucial for improving the pork industry. We previously created a Zinc finger-BED domain transcription factor (ZBED6)-deficient pig model which exhibited accelerated postnatal growth. Here, we evaluated the effect of ZBED6 on meat quality, flavor, nutritional value, safety and the mechanisms underlying meat production in pigs. Our results indicated that ZBED6 deficiency enlarges body size by enhancing feed efficiency. The results of carcass characteristics and meat quality measurements showed that ZBED6 deficiency enhances carcass lean percentage (46.49±0.62 % for WT vs. 52.70±0.56 % for ZBED6^-/-; P<0.001) and improves redness (12.39±0.42 for WT vs.14.53±0.59 for ZBED6^-/-; P=0.04) and reduces cooking loss (50.34±0.43% for WT vs.48.34±0.55% for ZBED6^-/-; P=0.04). Analysis of fatty acid and amino acid profiles showed that ZBED6 deficiency enhances both the nutritional value and flavor of pork. A comprehensive analysis utilizing RNA-seq, quantitative proteomics, and ChIP-seq identified the immunoglobulin superfamily containing leucine-rich repeat (ISLR) as a direct negative target of ZBED6. In C2C12 cells with knockdown of Zbed6, Islr expression is elevated, activating the canonical Wnt pathway and promoting myoblast differentiation and myotube formation, while knockdown of Islr significantly attenuated these effects. The subchronic oral toxicity study of ZBED6 deficiency pork in rat revealed no significant differences in daily clinical signs, body weight, feed intake, hematology, and serum biochemistry compares to wild-type pork. In summary, our study demonstrates the potential of ZBED6-deficient pigs as a valuable resource for the livestock and food industry, providing new insights into the mechanisms by which ZBED6 promotes muscle growth through the regulation of ISLR pathway.

Keywords: Carcass traits ISLR Meat quality Wnt signaling pathway Zinc finger BED domain containing protein 6

Online: 18 April 2025

Fund:

This research was supported by the National Key Research and Development Program of China (2021YFF1000602), the STI2030-Major Projects (2023ZD0404702), the Program for Shaanxi Science and Technology from Shaanxi Provincial Science and Technology Department (2023-CX-TD-57), and the National Natural Science Foundation of China (32400424).

About author: Huan Liu, huanliu150808@nwafu.edu.cn; Jiahe Huang, HuangJiaHeNLA@163.com; Junying Xiao, xiaojunying@nwafu.edu.cn; # Correspondence Jiangwei Wu, E-mail: wujiangwei@nwafu.edu.cn; Dengke Pan, E-mail: pandengke2002@163.com * These authors contributed equally to this work

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Huan Liu, Jiahe Huang, Junying Xiao, Ruirui Pan, Ruixin Zhang, Xiangyang Xing, Xiao Zhang, Bo Xia, Dengke Pan, and Jiangwei Wu. 2025. ZBED6 deficiency enhances porcine carcass traits and meat quality via ISLR-mediated Wnt signaling activation. Journal of Integrative Agriculture, Doi:10.1016/j.jia.2025.04.018

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