Interplay between mitophagy and substrate utilization in heart failure progression

项目总结 线粒体功能障碍和线粒体新陈代谢改变与发育有关。 心力衰竭(HF)。有丝分裂吞噬是一种特殊的自噬途径，介导溶酶体依赖 清除受损的线粒体，对线粒体质量控制至关重要。然而，我们目前 关于心脏中的有丝分裂及其如何与心肌代谢相关的知识有限。低于正常 在这种情况下，心脏主要依靠脂肪酸β氧化(粮农组织)来补充三磷酸腺苷的产生。相比之下，一个 心脏衰竭或肥大通常表现为粮农组织受损和对葡萄糖利用的依赖增加。尽管 在理解削弱粮农组织的监管计划方面取得了重大进展，目前尚不清楚如何转变 在心肌细胞中，底物的利用有助于有丝分裂的调节。因此，这项研究建议 重点阐述了心肌线粒体吞噬反应对心肌底物利用改变的功能意义 例如，在受损的粮农组织的背景下就会发生这种情况。这个项目的目的是描绘这部小说 有丝分裂和心肌底物利用之间的机制联系，以及确定 吞丝分裂是否代表了心脏病治疗的新机制和治疗靶点。 这些研究将由我们最近描述的监测活体心脏的mt-keima小鼠模型来促进 有丝分裂，以及一系列从遗传和药物上调节有丝分裂的创新试剂 流量。为了直接评估FAO在心脏中的作用，我们培育了心肌细胞特异性缺失的小鼠 CPT2(CPT2-CKO)，编码粮农组织所需的单一基因。我们已经证明了吞丝分裂的下降。 在粮农组织缺乏的心脏中出现心脏功能受损的先兆。我们的基因分析表明 心脏CPT2缺失损害PTEN诱导的PINK1信号通路，这是积极的 通过线粒体泛素化调节有丝分裂。通过调节USP30增强有丝分裂吞噬作用， 它介导了线粒体上的去泛素化这一反向反应，缓解了粮农组织的功能衰退 有缺陷的心脏。因此，在拟议研究的目标1中，我们的目标是定义 粮农组织受损对心脏有丝分裂的反应，以及对潜在的分子机制的剖析 将有丝分裂与心肌底物利用联系起来。在拟议研究的目标2中，我们将从基因上 并使用缺乏USP30的小鼠模型从药理上操纵USP30在心脏中的酶功能 或者用一种新型的USP30抑制剂治疗。我们将确定有害的心脏表型， 由心脏CPT2缺失或压力超负荷引起的，可以通过恢复至少部分逆转 通过抑制USP30抑制心肌细胞的有丝分裂。拟议研究的完成将产生关键的 对吞噬有丝分裂在正常心血管生理和病理条件中的作用的见解，并将 从根本上促进了我们对线粒体代谢和线粒体之间相互作用的理解 心脏中线粒体的质量控制。

PROJECT SUMMARY Mitochondrial dysfunction and altered mitochondrial metabolism have been implicated in the development of heart failure (HF). Mitophagy is a specialized autophagic pathway that mediates the lysosome-dependent clearance of damaged mitochondria, and is essential for mitochondrial quality control. However, our current knowledge regarding mitophagy in the heart and how it relates to myocardial metabolism is limited. Under normal conditions, the heart relies predominantly on fatty acid β-oxidation (FAO) to fuel ATP production. In contrast, a failing or hypertrophied heart usually shows impaired FAO and increased reliance on glucose utilization. Despite significant advancements in understanding the regulatory programs that attenuate FAO, it is unclear how shifts in myocardial substrate utilization contribute to the regulation of mitophagy. Therefore, this research proposal focuses on the functional significance of cardiac mitophagy in response to altered myocardial substrate utilization that occurs, for instance, in the setting of impaired FAO. The goal of this project is to delineate the novel mechanistic link between mitophagy and myocardial substrate utilization in the heart, as well as to determine whether mitophagy represents a novel mechanism and therapeutic target for the treatment of heart disease. These studies will be facilitated by our recently described mt-Keima mouse model to monitor in vivo cardiac mitophagy, as well as a set of innovative reagents to genetically and pharmacologically modulate mitophagic flux. To directly assess the role of FAO in the heart, we have generated mice with cardiomyocyte-specific deletion of CPT2 (CPT2-cKO), encoding a single gene required for FAO. We have demonstrated a decline in mitophagy precedes the development of impaired cardiac function in FAO-deficient hearts. Our genetic analyses suggest cardiac CPT2 deletion impairs the PTEN-induced putative kinase 1 (PINK1) signaling pathway, which positively regulates mitophagy through mitochondrial ubiquitination. Augmentation of mitophagy by modulating USP30, which mediates the reverse reaction, deubiquitination on mitochondria, mitigates the functional decline in FAO deficient hearts. Therefore, in Aim 1 of the proposed studies, our goal is to define the magnitude and Kinetics of cardiac mitophagy in response to impaired FAO, as well as to dissect the underlying molecular mechanisms connecting mitophagy to myocardial substrate utilization. In Aim 2 of the proposed studies, we will genetically and pharmacologically manipulate USP30 enzymatic function in the heart using mouse models that lack USP30 or are treated with a novel USP30 inhibitor. We will determine whether the detrimental cardiac phenotype, induced by cardiac CPT2 deletion or pressure overload, could be reversed, at least partially, by restoring mitophagy in cardiomyocytes via the inhibition of USP30. Completion of the proposed studies will produce critical insights into the role of mitophagy in normal cardiovascular physiology and in pathological conditions, and will fundamentally advance our understanding of the interaction between mitochondrial metabolism and mitochondrial quality control in the heart.