Mechanisms of Selective Autophagy

 描述(申请人提供)：选择性自噬巨型自噬的机制(称为自噬)是细胞通过高度调控的途径隔离胞液成分/细胞器的过程，该途径以溶酶体介导物的降解和大分子循环回到胞浆中而终止。虽然最初被认为是大量降解的非特定过程，但最近的工作揭示了几种形式的选择性自噬，它们在多种生物过程中发挥关键作用。各种类型的选择性自噬在多个水平上受到调控，使用一系列特定的自噬受体，这些受体识别不同的货物，以及与自噬小体相关并结合货物受体的不同ATG8蛋白家族。先前的工作表明，自噬通过铁蛋白(一种形成一种隔离游离铁的复合体的蛋白质)的降解来控制细胞铁水平；然而，这种活动背后的机制尚不清楚。通过先进的定量蛋白质组学，我们的实验室发现NCOA4是铁蛋白降解(铁蛋白吞噬)的自噬受体。此外，我们的初步数据表明，这一过程在几个水平上受到高度调控，包括铁对NCOA4的降解。这些发现为我们提供了前所未有的机会，机械地剖析吞铁蛋白的生化基础，以及阐明吞铁蛋白如何与有助于铁稳态和氧化应激控制的其他形式的选择性自噬(即有丝分裂吞噬)相结合。这一提议的首要假设是，各种形式的选择性自噬受到高度调控，以协调关键的细胞过程，如调节生物可利用的铁。在此背景下，我们提出了以下目标： 目的1.调节铁吞噬和整合到全球自噬途径的生化机制。这些研究将阐明在铁蛋白吞噬和其他形式的选择性自噬中，货物识别和招募对自噬小体的选择性的潜在机制。 目的2.探讨细胞应激时吞噬丝裂原和吞铁蛋白的协同作用。这些研究将阐明吞铁蛋白和有丝分裂吞噬对关键细胞功能的贡献。 目的3.通过对体内铁代谢的调控，阐明自噬在体内组织动态平衡中的重要性。这些研究将使用自噬以诱导的方式有条件地被抑制的小鼠模型，以及条件NCOA4基因敲除小鼠，以了解自噬，特别是铁素噬菌体通过控制生物可利用铁在红系发生中的作用。

 DESCRIPTION (provided by applicant): Mechanisms of Selective Autophagy Macroautophagy (referred to as autophagy) is a process whereby cells sequester cytosolic components/organelles via a highly regulated pathway that terminates with the lysosomal-mediated degradation of the cargo and the recycling of macromolecules back into the cytosol. While initially thought to be a non-specific process of bulk degradation, recent work has revealed several forms of selective autophagy that play key roles in multiple biological processes. The various types of selective autophagy are regulated on multiple levels, using a repertoire of specific autophagy receptors, which recognize distinct cargo as well as a diverse family of ATG8 proteins that associate with the autophagosome and bind cargo receptors. Prior work has implicated autophagy in the control of cellular iron levels through the degradation of ferritin (a protein that forms a complex which sequesters free iron); however, the mechanism underlying this activity remained unclear. Through advanced quantitative proteomics, our laboratories discovered NCOA4 as the autophagy receptor for ferritin degradation (ferritinophagy). Moreover, our preliminary data suggests that this process is highly regulated at several levels, including the degradation of NCOA4 in response to iron. These findings provide us with the unprecedented opportunity to mechanistically dissect the biochemical basis for ferritinophagy, as well as to elucidate how ferritinophagy is integrated with other forms of selective autophagy (i.e. mitophagy) that contribute to iron homeostasis and oxidative stress control. The overarching hypothesis of this proposal is that the various forms of selective autophagy are highly regulated to coordinate critical cellular processes such as the regulation of bioavailable iron. Against this backdrop, we propose the following Aims: AIM 1. BIOCHEMICAL MECHANISMS REGULATING FERRITINOPHAGY AND INTEGRATION INTO GLOBAL AUTOPHAGY PATHWAYS. These studies will elucidate mechanisms underlying the selectivity of cargo recognition and recruitment to autophagosomes in ferritinophagy and other forms of selective autophagy. AIM 2. TO EXPLORE THE COOPERATIVE FUNCTIONS OF MITOPHAGY AND FERRITINOPHAGY IN RESPONSE TO CELLULAR STRESSORS. These studies will elucidate the contributions of ferritinophagy and mitophagy to critical cellular functions. Aim 3. TO ELUCIDATE THE IMPORTANCE OF AUTOPHAGY IN TISSUE HOMEOSTASIS IN VIVO THROUGH THE CONTROL OF IRON METABOLISM. These studies will use mouse models where autophagy is conditionally inhibited in an inducible fashion as well as a conditional NCOA4 knockout mouse to understand the role of autophagy and specifically ferritinophagy in erythrogenesis through its control of bioavailable iron.