Exploring regulatory mechanisms of glyoxalase-1

项目摘要 甲基乙二醛(MG)是一种有效的细胞内糖基化试剂，可形成高级糖基化终产物。已形成 自发地从3-碳糖酵解中间体，MG迅速糖化蛋白质和核苷酸，损害 线粒体，并直接增加活性氧的产生，从而诱导促氧化状态和 类似衰老的状态。MG和相关的乙草酸酶酶防御系统正在成为关键 在老龄化和年龄相关疾病过程中的参与者。在生理条件下，MG会迅速解毒 乙草酸酶1(GLO1)。然而，当GLO1减弱时，MG通量增加，MG修饰的蛋白质 在细胞内和细胞外积累(称为二羰基胁迫)。二羰基应激促进血糖 不耐受、氧化应激和炎症。GLO1蛋白稳定性和酶活性的调控机制 骨骼肌组织是一种对葡萄糖代谢至关重要的组织，其活性还没有得到很好的研究，而且有一个关键的 需要了解GLO1降低在肥胖、老龄化和年龄背景下的功能后果- 相关疾病。GLO1对细胞功能至关重要，并受到许多翻译后修饰的影响 (PTMS)调节GLO1蛋白的稳定性和活性。我们的目标是建立健壮的翻译模型 描述GLO1被调控的机制，以更好地理解 减弱的GLO1。新的、最先进的翻译模式的产生将有助于加快 了解GLO1衰减和二羰基应激及其对骨骼肌健康的影响 无论是寿命还是健康寿命。我们的目标是建立GLO1缺失和GLO1缺失之间的功能相关性 GLO1的PTMS对人肌管的影响我们的方法是减弱GLO1并突变关键氨基酸 残基使用CRISPR基因编辑技术，再加上二羰基胁迫的措施。我们希望 确定GLO1调节失调和甲基乙二醛介导的损伤的一种新的肌肉特异性机制。这个 这项工作的圆满完成将对增进理解产生重要的积极影响，并 提供潜在的治疗靶点，以维持骨骼肌功能与衰老和年龄相关疾病。

PROJECT ABSTRACT Methylglyoxal (MG) is a potent intracellular glycating agent that forms advanced glycation endproducts. Formed spontaneously from 3-carbon glycolytic intermediates, MG rapidly glycates proteins and nucleotides, damages mitochondria and directly increases reactive oxygen species production; thus inducing a pro-oxidant state and senescent-like condition. MG and the related glyoxalase enzymatic defense system are emerging as critical players in aging and age-related disease processes. Under physiologic conditions MG is rapidly detoxified by glyoxalase 1 (GLO1). However, when GLO1 is attenuated, MG flux is increased and MG-modified proteins accumulate (termed dicarbonyl stress), both within and outside the cell. Dicarbonyl stress promotes glucose intolerance, oxidative stress and inflammation. The mechanisms regulating GLO1 protein stability and enzymatic activity in skeletal muscle tissue, a tissue critical to glucose metabolism, are not well studied and there is a critical need to understand the functional consequences of reduced GLO1 in the context of obesity, aging and age- related disease. GLO1 is critical to cellular function and subject to numerous posttranslational modifications (PTMs) that regulate GLO1 protein stability and activity. Our objective is to establish robust translational models to delineate the mechanisms by which GLO1 is regulated to better understand the functional consequences of attenuated GLO1. The generation of new, state-of-the-art translational models will help to accelerate the understanding of GLO1 attenuation and dicarbonyl stress and the implications for skeletal muscle health across both the life-span and health-span. We aim to establish the functional relevance of both GLO1 loss and the impact of PTMs of GLO1 in human myotubes. Our approach is to attenuate GLO1 and mutate critical amino acid residues using CRISPR gene editing technology, coupled with measures of dicarbonyl stress. We expect to identify a novel, muscle specific mechanism of GLO1 dysregulation and methylglyoxal-mediated damage. The successful completion of this work will have an important positive impact on advancing the understanding, and provide potential therapeutic targets, to maintain skeletal muscle function with aging and age-related disease.