O-GlcNAC糖基化修饰对冷暴露仔猪骨骼肌代谢的影响及其稳态重构机制

Cold damage is one of the toughest problem when people are facing ultra-cold weather, and also a challenge in stockbreeding development in north area. In our previous work, we have verified that O-GlcNAc as a stress sensor play an important role in the reaction of cold adaptation in animals. We propose to use neonatal piglets as experiment animal to select and identify for the key proteins of O-GlcNAcylation under the condition of cold stress, then construct the signal pathways and metabolic pathways for O-GlcNAcylation; Then we use OGT gene knockout mouse as a model animal to study the changes of signal pathway for regulating muscle metabolism in the condition of deficiency of OGT gene, and definite which is the crucial pathway for O-GlcNAc in the reaction of cold stress, and we also study the effect of O-GlcNAc for protein post-translational modification on the condition of deficiency of OGT gene. By the experiment of in vivo and in vitro combination, we also verify the signal pathway and protein post-translational modification mechanism for regulating muscle metabolism in the reaction of cold stress for muscle cells, and we will illuminate the homeostasis reestablishment mechanism for the body recover from imbalance state to physiology homeostasis. Our research aims to clarify the signal pathways of O-GlcNAc regulating muscle metabolism and its homeostasis reestablishment mechanism for protein post-translational modification on the reaction of cold exposure in neonatal pigs, and provide theoretical foundation to the elucidation of the physiology mechanisms of cold adaption and sustainable development of animal husbandry.

北方高寒地区冷害已成为人类面对极端严寒天气及发展畜牧业颇为棘手的问题之一。前期工作中，我们初步验证O-GlcNAc作为一种“应激感受器”在冷应激反应中发挥调节作用，但其机制尚不清楚。本研究拟通过仔猪在冷应激下骨骼肌中O-GlcNAc糖基化修饰关键蛋白的筛选，构建其参与冷应激调节相关的信号转导通路及其代谢途径；利用模式动物OGT基因敲除鼠，研究冷应激下OGT基因缺失时骨骼肌O-GlcNAc调控的信号通路的变化及对蛋白质O-GlcNAc糖基化修饰的影响；通过体外实验和体内实验相结合，明确调控骨骼肌细胞在冷应激反应中的信号调节机制及其在蛋白质翻译后修饰中的作用，阐明机体在经受冷应激后从失衡态到生理稳态的重构机制。研究结果可望阐明O-GlcNAc糖基化修饰对冷暴露仔猪骨骼肌代谢调控的信号途径及其蛋白质翻译后修饰的稳态重构机制，为揭示机体冷适应的分子生物学奥秘及畜牧业可持续发展奠定理论依据。

低温环境下骨骼肌代谢稳态调控一直是环境生理及代谢生理学的前言科学问题。前期工作中，我们初步证明O-GlcNAc糖基化修饰作为一种“应激感受器”在冷应激反应中发挥调节作用，但其机制尚不清楚。基于此，本项目以模式动物：骨骼肌OGT基因敲除鼠及仔猪为研究对象，通过体外实验和体内实验，对低温环境下O-GlcNAc糖基化修饰在小鼠及仔猪骨骼肌代谢调控稳态中的作用进行了一系列的研究。.首先我们对O-GlcNAc糖基化修饰在急性冷暴露下小鼠骨骼肌葡萄糖代谢稳态中的作用进行了初步探究。结果表明急性冷暴露可引起全身葡萄糖水平下降，胰岛素抵抗和腓肠肌组织中糖原含量及ATP水平降低；而急性冷暴露可诱导小鼠腓肠肌组织中整体O-GlcNAc修饰水平的变化，这可能是导致组织细胞凋亡减少、促进细胞存活和葡萄糖代谢调节的因素之一。不仅如此，本研究还发现急性冷暴露下，骨骼肌中OGT基因缺失可影响机体能量代谢和胰岛素敏感性；并且O-GlcNAc糖基化修饰可通过调节IRS-1/AKT信号通路来调控下游葡萄糖代谢相关蛋白，增强腓肠肌组织葡萄糖代谢及胰岛素敏感性。除此之外，本研究还发现急性冷暴露下小鼠腓肠肌中OGT介导RBM3蛋白表达上调，而RBM3的上调能够促进AKT磷酸化从而增强糖酵解和细胞保护；并且急性冷暴露条件下OGT介导的O-GlcNAc糖基化修饰调控NF-κB活性进而上调小鼠骨骼肌IL-6表达，进而促进骨骼肌与脂肪间的Crosstalk。最后本研究以仔猪为研究对象，系统证明了急性冷暴露条件下，O-GlcNAc糖基化修饰可通过影响仔猪腓肠肌组织中的葡萄糖代谢进而改善线粒体稳态实现细胞保护作用。.综上所述，本研究通过在骨骼肌OGT基因敲除鼠及仔猪体内的系统讨论，充分阐明了O-GlcNAc糖基化修饰对冷暴露仔猪骨骼肌代谢的调控作用及其相关途径，为揭示机体冷适应的分子生物学奥秘及畜牧业可持续发展奠定理论依据和潜在的干预靶点。