A model for elimination of defective mitochondrial genomes

摘要 这项拟议的研究的主要目标是揭示消除线粒体缺陷的机制。 基因组(MtDNA)通过遗传和表观遗传过程。线粒体DNA在动物细胞中的异质性 允许积累具有潜在复制优势的有缺陷的mtDNA。质量控制系统的功能 移除这种有害的线粒体基因组。我们发现携带大量缺失的丰度 线虫中的mtDNA，uaDf5，通常与野生型mtDNA稳定地保持在异质状态 (WT-mtDNA)随着成年年龄的增长而增加，这种增加的负担会传递给年长母亲的后代。 这种有缺陷的mtDNA在缺乏两种质量控制系统之一的突变体中也会升高：有丝分裂(粉红色- 1(-))或生殖系细胞程序性死亡(PCD；Ced-13(-))。然而，出乎意料的是，我们发现当两者都 PINK-1和CED-13功能被消除，相反的情况发生：uaDf5被快速且完全去除， 有效治愈动物的线粒体疾病。更令人震惊的是，它们的后代 PINK-1(-)或Ced-13(-)单一突变母亲的男性同时缺乏PINK-1和CED-13，也表现出快速 去除缺失的mtDNA，并且这种效果与不同的后代基因类型无关。这些 这些发现使我们假设，来自这些杂交的启动遗传事件(IGE)触发了一种有效的 区分并有效去除有缺陷的线粒体DNA的跨代表观遗传过程。有了这些 基础发现，我们将探讨这一惊人的线粒体DNA质量控制过程的机制。在AIM 1，我们将分析IGE触发的uaDf5移除对其年龄相关性积累的影响 分析其发展时机。我们将测试消除过程是否适用于其他mtDNA 并评估大小和序列位置对移除过程的影响。我们会调查的 吞丝分裂和PCD途径中的其他成分是否在激活去除方面显示出协同作用，并将测试 增强线粒体分裂的假说可能允许区分和消除uaDf5。在AIM 2、我们将分析mtdna质量明显的表观遗传性跨代传递的基础。 控制过程。我们将评估我们发现的WT-mtDNA戏剧性扩增的假设 发生在移除过程需要移除uaDf5且uaDf5是 触发这种放大所需的。我们将研究IgE是否拮抗线粒体UPR， 它可以保护uaDf5不被移除。我们将检验这一假设，即免疫球蛋白E是由父亲特异性地触发的 失去了PINK-1和CED-13。最后，我们将评估免疫球蛋白激活的表观遗传过程是否被传递 通过细胞核或细胞质/线粒体。这些研究将有助于我们理解 保持健康的线粒体基因组，这是一个对衰老、神经变性和 线粒体疾病。

SUMMARY The major objective of the proposed research is to unveil mechanisms that eliminate defective mitochondrial genomes (mtDNA) through genetic and epigenetic processes. The heteroplasmic state of mtDNA in animal cells allows accumulation of defective mtDNAs with potential replicative advantage. Quality control systems function to remove such deleterious mitochondrial genomes. We found that the abundance of a large deletion-bearing mtDNA in C. elegans, uaDf5, that is normally stably maintained in a heteroplasmic state with wild-type mtDNA (WT-mtDNA) increases with adult age and that this increased burden is passed onto offspring of older mothers. This defective mtDNA is also elevated in mutants lacking either of two quality control systems: mitophagy (pink- 1(-)) or germline programmed cell death (PCD; ced-13(-)). Unexpectedly, however, we found that when both pink-1 and ced-13 functions are eliminated, the opposite occurs: uaDf5 is rapidly and completely removed, effectively curing the animal of mitochondrial disease. Even more startling, descendants of crosses between pink-1(-) or ced-13(-) single mutant mothers with males lacking both PINK-1 and CED-13 also show rapid removal of the deleted mtDNA, and this effect occurs irrespective of the differing descendant genotypes. These findings lead us to hypothesize that an initiating genetic event (IGE) from these crosses triggers a potent transgenerational epigenetic process that discriminates and effectively removes defective mtDNA. With these foundational findings, we will investigate the mechanisms of this striking mtDNA quality control process. In Aim 1, we will analyze the effect of the IGE-triggered removal of uaDf5 on its age-dependent accumulation and analyze its developmental timing. We will test whether the elimination process is general to other mtDNA deletions and assess the impact of size and sequence location on the removal process. We will investigate whether other components in the mitophagy and PCD pathways show synergy in activating removal and will test the hypothesis that enhanced mitochondrial fission may allow for discrimination and elimination of uaDf5. In Aim 2, we will analyze the basis for the apparently epigenetic transgenerational transmission of the mtDNA quality control process. We will evaluate the hypothesis that the dramatic amplification of WT-mtDNA that we found occurs prior to the generations in which uaDf5 is removed is required for the removal process and that uaDf5 is required to trigger this amplification. We will investigate whether the IGE antagonizes the mitochondrial UPR, which protects uaDf5 from removal. We will test the hypothesis that the IGE is triggered specifically by paternal loss of PINK-1 and CED-13. Finally, we will assess whether the IGE-activated epigenetic process is transmitted through the nucleus or cytoplasm/mitochondria. These studies will contribute to our understanding of how a healthy mitochondrial genome is maintained, a problem of premier importance to aging, neurodegeneration, and mitochondrial disease.