A transmembrane Rab GTPase accelerating protein targeted to peroxisomes

项目摘要 了解细胞如何适应压力和修复损伤是细胞生物学的最高优先事项之一， 健康研究。内质网（ER）是用于构建和分泌细胞的脂质和蛋白质的关键来源。 维持细胞中的几个细胞器，并在响应刺激时，它能够将资源引导到一个 各种运输途径。信号蛋白Ypt 1/Rab 1是Rab GT3（Rab）， 在ER如何指导资源和执行受损细胞器的质量控制方面发挥重要作用，包括 线粒体和过氧化物酶体。我们的长期目标是了解脂质和蛋白质 货物被路由和重新路由到特定的行程，以响应细胞的刺激和压力，这将 为制定针对疾病的有针对性的治疗干预措施提供信息。我们发现 GT3蛋白加速蛋白Gyp 8是一种进化上保守但知之甚少的GT3负调控因子， Ypt 1/Rab 1信号传导定位于ER、过氧化物酶体和线粒体，并影响Ypt 1信号传导。我们 中心假设是Gyp 8在早期分泌途径中起调节Ypt 1/Rab 1信号传导的作用 (ER)和非分泌膜（过氧化物酶体和线粒体），经常受到损害 通过Ypt 1/Rab 1依赖的选择性自噬进行质量控制。此外，由于Gyp 8定位于细胞器， 通常在必需的代谢途径中与交换材料连接/对接，我们预测Gyp 8 调节Rab-dependent tethering相互作用，特别是在专门用于脂质储存的细胞器中， 新陈代谢.为了验证我们的中心假设，并推进对几种膜生物发生的理解， 我们将追求这些特定的目标：1）识别基因内和基因外因素 2）确定由Gyp 8的亚细胞行程和活性调节的靶Rab GT3; Gyp 8在分泌途径中的作用;以及3）确定Gyp 8在调节过氧化物酶体和线粒体中的作用 动力学拟议的研究是创新的重点领域和技术方法。而 Ypt 1/Rab 1信号转导控制着细胞健康所必需的多种转运途径，了解其中的 并且当信号必须被终止以完成其每一个角色时是特别不完整的。的 实验计划结合了金标准生物化学和成像技术在细胞器和囊泡 转运（酶偶联动力学转运测定和3D电子断层扫描）与系统生物学 方法（合成基因阵列和亲和捕获质谱蛋白质组学）。拟议研究 是重要的，因为定义Ypt 1/Rab 1是如何调节的，以支持和行使ER的质量控制， 线粒体和过氧化物酶体是理解心血管、神经退行性疾病 和代谢紊乱。

Project Summary Understanding how cells adapt to stress and repair damage is one of the highest priorities in cell biology and health research. The endoplasmic reticulum (ER) is a key source of lipid and proteins for building and maintaining several organelles in cells, and in response to stimuli it is capable of directing resources into an assortment of transport pathways. The signaling protein Ypt1/Rab1 is a Rab GTPase (Rab) that plays essential roles in how the ER directs resources and executes quality control of damaged organelles including mitochondria and peroxisomes. Our long-term goal is to understand the mechanisms for how lipid and protein cargos are routed and re-routed to specific itineraries in response to cellular stimuli and stresses, which will inform development of targeted therapeutic interventions to address diseases. We have found that the GTPase accelerating protein Gyp8, an evolutionarily conserved but poorly understood negative regulator of Ypt1/Rab1 signaling, localizes to the ER, peroxisomes and mitochondria and impinges on Ypt1 signaling. Our central hypotheses are that Gyp8 functions to modulate Ypt1/Rab1 signaling in the early secretory pathway (ER) and at non-secretory membranes (peroxisomes and mitochondria) that are subject to frequent damage and quality control via Ypt1/Rab1-dependent selective autophagy. Also, since Gyp8 localizes to organelles that commonly tether/dock to exchange materials in essential metabolic pathways, we predict that Gyp8 regulates Rab-dependent tethering interactions, particularly among organelles specialized for lipid storage and metabolism. To test our central hypotheses and advance understanding of several membrane biogenesis pathways that originate at the ER, we will pursue these specific aims: 1) Identify intra- and extra-genic factors that control the subcellular itinerary and activity of Gyp8; 2) Determine target Rab GTPase(s) regulated by Gyp8 in the secretory pathway; and 3) Define the role of Gyp8 in regulating peroxisomal and mitochondrial dynamics. The proposed research is innovative both for area of focus and technical approach. While Ypt1/Rab1 signaling controls multiple transport pathways essential to cellular health, understanding of where and when signal must be terminated to accomplish each of its roles is particularly incomplete. The experimental plan combines gold standard biochemical and imaging techniques in organelle and vesicular transport (enzyme-coupled kinetic transport assays and 3D electron tomography) with systems biology approaches (synthetic gene array and affinity capture mass spectrometry proteomics). The proposed research is significant because defining how Ypt1/Rab1 is regulated to support and exercise quality control of ER, mitochondria and peroxisomes is foundational to understanding aspects of cardiovascular, neurodegenerative and metabolic disorders.