Nuclear-mitochondrial co-regulation during mitochondrial biogenesis

项目总结/摘要 线粒体氧化磷酸化（OXPHOS）机制的组装和维护缺陷 导致一系列退行性疾病，包括糖尿病、癌症和神经退行性疾病。 OXPHOS复合物在细胞核和线粒体基因组上都有编码，因此它们的生物合成 需要跨基因组的基因调控机制的精确协调。为此，线粒体 生物发生和应激反应程序涉及核转录的同时上调， 编码OXPHOS基因和线粒体基因表达因子。这些转录程序是 被认为促进核-线粒体平衡，在此，线粒体编码的OXPHOS亚基组装 与它们的核编码对应物的化学计量比。我们最近在酵母中观察到 酿酒酵母细胞核编码和细胞质编码的OXPHOS亚基的转录调控是 在碳源适应过程中不协调。相反，细胞抑制了细胞的翻译调节， OXPHOS亚基跨隔室。同步翻译是否是一种广泛的反应， 未知该提案的目标是确定线粒体和核基因组如何共同调节， 特别是在蛋白质合成期间，在整个线粒体生物发生和应激反应程序中。作为 作为我们最近在酵母中工作的延伸，Aim 1将研究同步翻译程序是否 发生在一系列环境和线粒体应激适应程序中。我们还将确定 如何通过确定关键线粒体翻译的作用来进行同步翻译调节 OXPHOS基因在碳源适应过程中的动态调节。为此将 使用我们的线粒体核糖体分析方法和细胞溶质核糖体分析来测量蛋白质 合成和RNA-seq以测量整体转录。在目标2和3中，我们将研究扩展到人类 细胞通过重新设计核糖体谱，以强大地捕获人类线粒体翻译。测试 方法，我们将研究如何推定的翻译激活剂，TACO 1，影响人类线粒体 翻译.我们将研究急性线粒体应激诱导的核-线粒体协同调节， 化学抑制OXPHOS复合物，模拟疾病过程中的OXPHOS功能障碍。最后我们 将研究iPS体外分化过程中线粒体和核基因表达程序 细胞到心肌细胞，当广泛的线粒体生物发生。确定法规和 核-线粒体协同调节的程度将为理解如何 在疾病状态中发生OXPHOS亚基的不平衡产生。

PROJECT SUMMARY/ABSTRACT Defects in the assembly and maintenance of mitochondrial oxidative phosphorylation (OXPHOS) machinery lead to a range of degenerative illnesses, including diabetes, cancer, and neurodegenerative diseases. OXPHOS complexes are encoded on both the nuclear and mitochondrial genomes, so their biogenesis requires the precise coordination of gene regulatory mechanisms across genomes. To this end, mitochondrial biogenesis and stress response programs involve the simultaneous transcriptional upregulation of nuclear- encoded OXPHOS genes and mitochondrial gene expression factors. These transcriptional programs are thought to facilitate nuclear-mitochondrial balance, where mitochondrial-encoded OXPHOS subunits assemble in stoichiometric ratios with their nuclear-encoded counterparts. We recently observed in Saccharomyces cerevisiae that transcription regulation of nuclear-encoded and mitochondrial-encoded OXPHOS subunits are not coordinated during carbon source adaptation. Instead, the cell synchronizes the translational regulation of OXPHOS subunits across compartments. Whether synchronized translation is a widespread response remains unknown. The goal of this proposal is to determine how mitochondrial and nuclear genomes are co-regulated, particularly during protein synthesis, throughout mitochondrial biogenesis and stress response programs. As an extension of our recent work in yeast, Aim 1 will investigate whether synchronous translation programs occur across a range of environmental and mitochondrial stress adaptation programs. We will also determine how the synchronous translation regulation occurs by determining the role of key mitochondrial translation regulators in the dynamic regulation of OXPHOS genes during carbon source adaptation. This will be done using our mitochondrial ribosome profiling approach and cytosolic ribosome profiling to measure protein synthesis and RNA-seq to measure global transcription. In Aims 2 and 3, we extend our studies to human cells through re-engineering ribosome profiling to robustly capture human mitochondrial translation. To test the approach, we will investigate how a putative translation activator, TACO1, impacts human mitochondrial translation. We will investigate nuclear-mitochondrial co-regulation after acute mitochondrial stress induced by chemical inhibition of OXPHOS complexes, mimicking OXPHOS dysfunction in disease processes. Finally, we will investigate mitochondrial and nuclear gene expression programs during the in vitro differentiation of iPS cells to cardiomyocytes, when extensive mitochondrial biogenesis occurs. Determining the regulation and the extent of nuclear-mitochondrial co-regulation will provide critical insight towards understanding how imbalanced production of OXPHOS subunits transpires in disease states.