Abstract
摘要:細胞自噬 (autophagy) 是真核細胞在適應生理壓力時的重要機制。細胞經由形成雙層膜的細胞自噬體 (autophagosome),將細胞質中巨分子物質送入濾泡(vacuole)或溶小體 (lysosome) 中,分解後釋出的基本成分可用於能量代謝或合成維持生命所需的新分子。早期的研究發現,缺乏養分的生理壓力是誘發非選擇性細胞自噬作用最有效的方法,因此過去一直認為細胞自噬不具有選擇被分解物的能力,而其功能則僅限於維持能量的恆定。然而,近年來研究則發現,在腫瘤與受組織退化疾病影響的細胞中,常伴隨有異常的細胞自噬現象,且抑制細胞自噬作用,會造成被泛素 (ubiquitin) 修飾過的蛋白質沉澱累積在細胞質中,顯示細胞自噬可選擇性去除異常的巨分子沉澱物及胞器,甚至促成細胞死亡,以避免這些物質干擾正常生理功能。因此,研究選擇性細胞自噬的分子篩選機制,不但有助於認識細胞自噬在疾病發生中所扮演的角色,更可能在將來用以發展新的療法。本計畫預定以三年時間,研究出芽酵母菌BAR-domain蛋白Atg20與Atg24,調控選擇性細胞自噬的分子機制。我們過去的實驗發現,細胞自噬蛋白Atg11經由與被運送物受體(cargo receptor)接合,協助選擇性細胞自噬的被運送物誘發細胞自噬體形成。然而,Atg11執行此功能的分子機轉仍有待進一步的研究確認。最近一年的實驗結果則顯示,Atg11會誘使二個帶有BAR-domain的蛋白Atg20與Atg24集中到細胞自噬體形成構造上,此過程為形成選擇性細胞自噬體不可或缺的條件;另外值得注意的是,Atg20與Atg24也會在內噬體 (endosome)處參與篩選被回收的細胞膜蛋白,因此我們推測,細胞自噬調控系統可能藉由Atg20以及Atg24的作用,將細胞的內膜運輸系統導向形成細胞自噬體。我們希望經由分析Atg20與Atg24在調控細胞自噬過程中的角色,讓我們更加瞭解選擇性細胞自噬作用在早期過程中受誘導發生的機制。
Abstract: Autophagy is a stress response mechanism that delivers cytosolic materials to the vacuole/lysosome for degradation and recycling. Because autophagy is highly activated by nutrient-starvation stresses, it had been considered simply as a non-specific catabolic pathway for a long time. Recent studies, however, have uncovered intimate relationships between selective autophagy activities and human diseases, such as pathogen infections and tissue-degenerative diseases. Based on results of animal model studies, autophagy is also proposed as a mechanism to prevent aging by eliminating dysfunctional mitochondria. All these newly reported autophagy functions rely on efficient signaling mechanisms to recognize and segregate specific cargoes for degradation, which makes the selective autophagy regulatory machinery a potential target for therapeutic manipulations. Working on a suitable model system should accelerate our studies on the molecular mechanisms of selective autophagic cargo-induced signaling processes. Several selective autophagic cargoes have been uncovered in the budding yeast Saccharomyces cerevisiae. Among them, the precursors of aminopeptidase I enzyme (prApe1) are delivered directly from cytosol to the vacuole through the actions of autophagy proteins under nutrient-rich growth condition. This makes prApe1 the only model for the study of the induction signaling of selective autophagy pathways in nutrient-rich environment. Because the other types of selective autophagy pathways are stimulated by nutrient-starvation conditions, which also induce non-selective autophagy activities, this makes the attribution of observed molecular events for selective or non-selective autophagy regulation difficult. We propose to study prApe1-stimulated selective autophagy regulation in this three-year proposal.
Atg19 is the sorting receptor for prApe1. Atg11 regulates vacuolar delivery of selective autophagic cargoes, including prApe1, peroxisomes, and mitochondria, revealing its adaptor function. Atg11 is proposed to perform this function by targeting cargo to the site of vesicle formation and serving as a scaffold protein for the construction of the phagophore assembly site (PAS), where autophagosomes eventually form. As a consequence, Atg11 is at a position to relay cargo-loading signals to vesicle formation machinery, but how this task is achieved is not clear. Atg11 is known to interact with many proteins. Among them, two yeast BAR-domain proteins, Atg20 and Atg24, are recruited by Atg11 to the PAS. Atg20 and Atg24 also contain a conserved phosphatidylinositol 3-phosphate-binding phox domain (PX domain), which is necessary for their localization and functions at the PAS. Interestingly, Atg20 and Atg24 are also found regulating membrane trafficking at the endosomes. Whether these two membrane trafficking regulatory functions of Atg20 and Atg24 are correlated have not been carefully examined. We propose to characterize the mechanisms underlying selective autophagy regulation by the BAR-domain protein Atg20 and Atg24 in molecular details.
Keyword(s)
選擇性細胞自噬
細胞膜運輸
分子篩選機制
出芽酵母菌
神經退化疾病
腫瘤
細胞死亡
selective autophagy
membrane trafficking
cargo-sorting mechanisms
Saccharomyces cerevisiae
neurodegeneration diseases
cancer
programmed cell death