The role of protein O-linked N-Acetylglucosamine in regulating cardiac physiology

O-连接N-乙酰氨基葡萄糖对蛋白质丝氨酸和苏氨酸残基的翻译后修饰 （O-GlcNAc）越来越被认为与磷酸化一样丰富，并发挥类似的作用。 在调节多种细胞功能，包括细胞周期，转录，蛋白质降解， 线粒体功能、自噬、昼夜节律和细胞存活。己糖胺生物合成的激活 调节蛋白质O-GlcNAc化的途径与两种C. 线虫和老鼠在心脏中，O-GlcNAc稳态的失调与不同的疾病有关。 过程包括心脏肥大、心力衰竭、糖尿病的不良影响和年龄相关变化 在心肌细胞中。另一方面，O-GlcNAc水平的急剧增加促进了 心肌细胞对缺血/再灌注损伤的反应。我们已经证明，O-GlcNAc酰化调节 这些生理过程是维持健康心肌细胞的关键。我们的初步数据还显示， 禁食增加了心脏中的O-GlcNAc水平，强烈支持蛋白质O- GlcNAc酰化。尽管O-GlcNAc与心脏疾病相关，但我们对O-GlcNAc的基本调节机制的了解， 在正常健康心脏的机制仍然有限，有越来越多的证据表明，O-GlcNAc循环 有助于调节健康心脏的正常生理过程。缺乏对 O-GlcNAc如何调节正常心肌细胞功能的基础生物学代表了一个主要的空白 并限制了在治疗心脏疾病中调节O-GlcNAc体内平衡的潜力。 疾病因此，我们认为，迫切需要更好地了解 O-GlcNAc对心肌细胞功能的影响。因此，我们将测试蛋白质O- GlcNAc酰化在正常心肌细胞稳态中起着不可或缺的作用， 心肌细胞对生理刺激的恢复力。我们将用两个具体的例子来验证这个假设。 目的：1）确定O-GlcNAc水平的变化如何影响心脏的分子和细胞反应 2）确定生理刺激如何影响OGT的时间和细胞内定位 和OGA和O-GlcNAc。我们将使用诱导型心肌细胞特异性小鼠模型来增加或减少 心脏中的O-GlcNAc水平、基于遗传的O-GlcNAc FRET传感器和荧光标记的OGT和 OGA以确定响应于生理刺激而发生的O-GlcNAc的时空动态。的 成功完成拟议的研究将产生关于O-GlcNAc在以下方面作用的重要新信息： 调节正常心脏生理学，包括其在维持心肌细胞弹性中的作用。本申请 通过专注于提高我们对心脏糖组的基础知识，直接响应PA-19-049 以及调节心血管生物学的能力。这些发现将奠定基础，将有助于进一步我们的 了解心脏健康和恢复力的信号通路。

The post-translational modification of serine and threonine residues on proteins by O-linked N-acetylglucosamine (O-GlcNAc), is increasingly recognized as being as abundant as phosphorylation and playing a similarly important role in regulating diverse cell functions including cell cycle, transcription, protein degradation, mitochondrial function, autophagy, circadian rhythm, and cell survival. Activation of the hexosamine biosynthesis pathway, which regulates protein O-GlcNAcylation has been associated with increased lifespan in both C. elegans and mice. In the heart dysregulation in O-GlcNAc homeostasis has been linked to different disease processes including cardiac hypertrophy, heart failure, the adverse effects of diabetes and age-related changes in cardiomyocytes. On the other hand, acute increases in O-GlcNAc levels promotes resilience of cardiomyocytes in response to ischemia/reperfusion injury. We have shown that O-GlcNAcylation regulates physiological processes that are key to maintaining healthy cardiomyocytes. Our preliminary data also show that fasting increases O-GlcNAc levels in the heart, strongly supporting a physiological role for protein O- GlcNAcylation. Despite the association of O-GlcNAc with cardiac disease our knowledge of the basic regulatory mechanisms in the normal healthy heart remains limited, there is growing evidence that O-GlcNAc cycling contributes to the regulation of normal physiological processes in a healthy heart. A lack of understanding of the fundamental biology underlying how O-GlcNAc regulates normal cardiomyocyte function represents a major gap in our knowledge and limits the potential for modulation of O-GlcNAc homeostasis in the treatment of cardiac disease. Consequently, we believe that there is an urgent need to better understand the basic biology underlying the effects of O-GlcNAc on cardiomyocyte function. Therefore, we will test the hypothesis that protein O- GlcNAcylation plays an integral role in normal cardiomyocyte homeostasis contributing to cardiomyocyte resilience in response to physiological stimuli. We will test this hypothesis with two specific aims: 1) Determine how changes in O-GlcNAc levels influence the molecular and cellular responses of the heart to fasting; and 2) Determine how physiological stimuli influence the temporal and intracellular localization of OGT and OGA, and O-GlcNAc. We will use inducible cardiomyocyte specific mouse models to increase or decrease O-GlcNAc levels in the heart, genetically based O-GlcNAc FRET sensors, and fluorescently tagged OGT and OGA to determine the spatiotemporal dynamics of O-GlcNAc that occur in response to physiological stimuli. The successful completion of the proposed studies will yield significant new information on the role of O-GlcNAc in regulating normal cardiac physiology including its role in maintaining cardiomyocyte resilience. This application is directly responsive to PA-19-049 by focusing on improving our fundamental knowledge of the cardiac glycome and its ability to regulate cardiovascular biology. These findings will lay the foundation that will help further our understanding of this signaling pathway in cardiac health and resilience.