Characterization of a novel autophagy pathway

 描述(申请人提供)：程序性细胞死亡在动物发育过程中起着重要作用，这一过程中的缺陷会导致多种人类疾病，包括癌症、神经退化和自身免疫。细胞凋亡和自噬细胞死亡是发生在发育过程中的两种最主要的细胞程序性死亡形态。对细胞凋亡的调控相对了解较多，但对介导自噬程序性细胞死亡的机制知之甚少。我们正在研究类固醇激活的果蝇自噬细胞死亡，并使用幼虫肠道的中肠作为模型。类固醇的增加会触发一种基因程序，从而激活中肠细胞死亡。这些发育调节的细胞死亡不依赖于包括半胱氨酸蛋白酶在内的凋亡基因，它们具有自噬细胞死亡所致细胞的形态。值得注意的是，自噬(ATG)基因是中肠降解所必需的，在那里它们调节程序化的细胞大小缩小。虽然酵母中巨型自噬(自噬)的功能和调控知道得很多，但对体内动物细胞中调节这一过程的机制知之甚少，对自噬在细胞死亡过程中的作用也知之甚少。人们一直认为控制自噬的机制在酵母和人类之间是相同的。我们的假设是，自噬在多细胞生物体中的细胞特异性使用涉及到以前未知的调控机制，这些机制与核心自噬途径整合在一起。为了支持这一假说，我们发现由Atg7和Atg3编码的保守的E1和E2酶不是苍蝇中肠自噬和降解所必需的，而这些基因是饥饿触发的果蝇自噬所必需的。相比之下，中肠细胞的自噬依赖于Uba1，即用于泛素化的E1。这些和其他数据表明，我们已经发现了一种新的机制，泛素通过它来调节ATG7和ATG3非依赖的自噬，我们的目标是表征在中肠细胞死亡过程中控制自噬的分子机制。在这里，我们建议：(1)研究Atg8在中肠自噬和细胞尺寸缩小中的作用，(2)确定泛素结合蛋白和Parkin底物在自噬中的作用，以及(3)鉴定清除线粒体和自噬所需的新基因。最近自噬与神经退行性疾病和癌症的关联表明，研究自噬在程序性细胞死亡中的作用是很重要的。

 DESCRIPTION (provided by applicant): Programmed cell death plays an important role during animal development, and defects in this process result in a variety of human disorders including cancer, neurodegeneration and autoimmunity. Apoptosis and autophagic cell death are the two most prominent morphological forms of programmed cell death that occur during development. The regulation of apoptosis is relatively well understood, but little is known about the mechanisms that mediate autophagic programmed cell death. We are studying steroid-activated autophagic cell death in Drosophila, and are using the midgut of the larval intestine as a model. An increase in steroid triggers a genetic program that activates midgut cell death. These developmentally-regulated cell deaths do not depend on apoptosis genes, including caspase proteases, and they possess the morphology of cells that die by autophagic cell death. Significantly, autophagy (Atg) genes are required for midgut degradation where they regulate programmed cell size reduction. While much is known about the function and regulation of macro-autophagy (autophagy) in yeast, less is known about the mechanisms that regulate this process in animal cells in vivo, and little is known about the function of autophagy during cell death. It has been assumed that the mechanisms controlling autophagy are identical between yeast and humans. Our hypothesis is that the cell-specific use of autophagy in multicellular organisms involves previously unrecognized regulatory mechanisms that integrate with core autophagy pathways. In support of this hypothesis, we have discovered that the conserved E1 and E2 enzymes encoded by Atg7and Atg3 are not required for autophagy and degradation of the fly midgut, while these genes are required for starvation-triggered autophagy in flies. By contrast, autophagy in midgut cells depends on Uba1, the E1 used for ubiquitination. These and other data indicate that we have discovered a novel mechanism by which ubiquitin regulates Atg7 and Atg3-independent autophagy, and our goal is to characterize molecular mechanisms that control autophagy during midgut cell death. Here we propose to: (1) investigate the role of Atg8 in midgut autophagy and cell size reduction, (2) determine the role of ubiquitin-binding proteins and Parkin substrates in autophagy, and (3) characterize new genes that are required for clearance of mitochondria and autophagy. The recent association of autophagy with neurodegenerative disorders and cancer indicates the importance of investigating the understudied role of autophagy during programmed cell death.