ADP: A Master Regulator for Bioenergetics and Ca2+/ROS Signaling in Heart

描述(由申请者提供)：心力衰竭是一个严重的医学问题：在美国有超过500万人受到影响。慢性心力衰竭的原因是多方面的，包括生物能量缺乏、钙超载和氧化应激。ADP是ATP形成的关键底物。此外，ADP对线粒体通透性转换孔(MPTP)的开放有很强的抑制作用。尽管MPTP的分子同一性仍未解决，但有两个不同的概念脱颖而出。一种认为MPTP不需要腺嘌呤核苷酸转位酶(ANT)，而另一种认为MPTP是一个多蛋白质复合体，ANT和线粒体内的亲环素-D(Cyp-D)是MPTP的关键成分。有趣的是，ADP与ANT的结合有助于心磷脂稳定呼吸链超复合体，从而提高ATP的生成效率。心磷脂的氧化破坏了这些超复合体的稳定，并与衰老、糖尿病心肌病、缺血再灌注损伤和心力衰竭相关的线粒体功能障碍有关。此外，ADP抑制无机磷PI的外流，无机磷PI是线粒体形成钙-PI复合体的关键钙缓冲物质。最后，线粒体裂变蛋白DLP1的激活导致线粒体分裂、ROS产生和MPTP开放。综上所述，这些结果使我们推测，ADP与ANT的结合在线粒体功能中起着两个基本作用：通过稳定心磷脂-ETC复合体来提高ATP的生成效率，以及通过减少ROS的产生和DLP1的激活来抑制MPTP。从生理上讲，ADP介导的MPTP抑制使线粒体过量的ROS产生和线粒体钙离子释放最小化，以优化兴奋-收缩-代谢(ECM)偶联的有效性。病理上，这种ADP调节机制的缺陷导致能量衰竭、氧化应激和钙离子失调，从而增加心脏对损伤的易感性。我们提出了两个特定的目标：特定的目标1：确定ADP抑制MPTP的机制。假设：ADP稳定心磷脂的完整性，抑制线粒体中DLP1的活性，从而防止Cyp-D依赖的MPTP开放。此外，ADP通过抑制MPTP减少ROS的产生，从而最大限度地减少ROS对MPTP的反馈激活。具体目的2：评价ADP在心脏保护中的作用。假设：疾病模型心脏中的MPTP对钙离子诱导的开放表现出更高的敏感性，这是因为它容易受到包括氧化应激、高DLP1活性和/或降低的钙缓冲能力在内的首次打击应激。维护最佳矩阵ADP水平可以缓解这种增加的MPTP漏洞。代谢信号和钙/氧化还原/细胞死亡信号之间相互作用的紊乱是包括心脏疾病在内的疾病发病机制中的基础。这一应用中的探索性和高风险想法如果得到验证，将在广泛的疾病机制和治疗方面开辟新的天地。 公共卫生相关性：不能以三磷酸腺苷的形式提供足够的细胞能量会导致许多人类疾病，包括心脏病发作和中风、神经退行性疾病、糖尿病和衰老。这项拟议的研究将探索ADP作为制造ATP的底物和氧化应激抑制物的双重作用。我们的目标是，不仅阐明ADP调节心肌细胞生死的基本机制，而且开发可能的治疗手段来治疗这些衰弱的疾病。

DESCRIPTION (provided by applicant): Heart failure is a serious medical problem: over 5 million people in the US affected. The cause for chronic heart failure is multifaceted and includes bioenergetic deficiency, Ca2+ overload, and oxidative stress. ADP is the key substrate for ATP formation. Moreover, ADP is a potent inhibitor for the opening of mitochondrial permeability transition pore (mPTP). Although the molecular identity of mPTP is still unsolved, 2 different concepts stand out. One indicates that adenine nucleotide translocase (ANT) is not required for mPTP, whereas the other depicts mPTP as a multi-protein complex, the ANT and the mitochondrial peptidyl-prolyl cis-trans isomerase known as cyclophilin-D (Cyp-D), are the key components. Interestingly, binding of ADP to ANT facilitates cardiolipin to stabilize respiratory chain supercomplexes so that the efficiency of ATP generation is enhanced. Oxidation of cardiolipin destabilizes these supercomplexes and has been linked to mitochondrial dysfunction associated with aging, diabetic cardiomyopathy, ischemia-reperfusion injury, and heart failure. Furthermore, ADP inhibits efflux of inorganic phosphate Pi, the key mitochondrial Ca2+ buffer that forms Ca2+-Pi complex. Finally, activation of the mitochondrial fission protein, DLP1 causes mitochondrial fission, ROS generation, and mPTP opening. Taken together, these results lead us to hypothesize that "binding of ADP to ANT serves two fundamental roles in mitochondrial function: enhancing ATP generation efficiency by stabilizing cardiolipin-ETC complexes and inhibiting mPTP by decreasing ROS generation and DLP1 activation. Physiologically, ADP- mediated mPTP inhibition minimizes excessive mitochondrial ROS generation and Ca2+ release from mitochondria in order to optimize the effectiveness of excitation-contraction-metabolism (ECM) coupling. Pathologically, defects of this ADP regulatory mechanism lead to energetic failure, oxidative stress, and Ca2+ dysregulation that enhance cardiac vulnerability to injury". We propose 2 Specific Aims: Specific Aim 1: To determine the mechanisms for ADP inhibition of mPTP. Hypothesis: ADP stabilizes cardiolipin integrity and inhibits DLP1 activity in the mitochondria and thus protects against Cyp-D-independent mPTP opening. Moreover, ADP decreases ROS generation via mPTP inhibition, and thus minimizes feedback activation of mPTP by ROS. Specific Aim 2: To assess the role of ADP in cardiac protection. Hypothesis: The mPTP in hearts of diseased models exhibits increased sensitivity to Ca2+-induced opening due to its predisposition to the first hit stresses including oxidative stress, high DLP1 activity, and/or diminished Ca2+ buffering capacity. Maintenance of optimal matrix ADP levels alleviates this increased mPTP vulnerability. Disturbances in the interaction between metabolic signaling and Ca2+/redox/cell death signaling are fundamental in disease pathogenesis including cardiac diseases. The exploratory and high-risk ideas in this application, if validated, will break new ground in a wide spectrum of disease mechanisms and treatments. PUBLIC HEALTH RELEVANCE: Failure to provide sufficient cellular energy in form of ATP can cause numerous human diseases including heart attacks and strokes, neurodegenerative diseases, diabetes, and aging. The proposed research will explore the dual role of ADP in serving as a substrate for making ATP and an inhibitor for oxidative stress. It is our objective, not only to elucidate the fundamental mechanisms how ADP regulates the life and death of heart cells, but also to develop possible therapeutic means for treating these debilitating disorders.