Fodor E, Sigmond T, Ari E, Lengyel K, Takács-Vellai K, Varga M, Vellai T
Eötvös Loránd University, Budapest, Hungary.
Eötvös Loránd University, Budapest, Hungary.
Methods Enzymol. 2017;588:467-496. doi: 10.1016/bs.mie.2016.10.028. Epub 2016 Dec 16.
Autophagy (cellular self-eating) is a highly regulated degradation process of the eukaryotic cell during which parts of the cytoplasm are delivered into, and broken down within, the lysosomal compartment. The process serves as a main route for the elimination of superfluous and damaged cellular constituents, thereby mediating macromolecular and organellar turnover. In addition to maintaining cellular homeostasis, autophagy is involved in various other cellular and developmental processes by degrading specific regulatory proteins, and contributing to the clearance of intracellular pathogens. The physiological roles and pathological involvement of autophagy can be effectively studied in divergent eukaryotic model systems ranging from yeast to mice. Such a tractable animal modelapplied only recently for autophagy researchis the zebrafish Danio rerio, which also facilitates the analysis of more specific biological processes such as tissue regeneration. In this chapter, we overview the main methods and tools that are used to monitor autophagic structures and to assay autophagic responses in this vertebrate organism. We place emphasis on genetic (functional) approaches applied for exploring novel cellular and developmental roles of the autophagic process.
自噬(细胞自噬)是真核细胞中一种高度调控的降解过程,在此过程中,部分细胞质被输送到溶酶体区室并在其中被分解。该过程是清除多余和受损细胞成分的主要途径,从而介导大分子和细胞器的更新。除了维持细胞内稳态外,自噬还通过降解特定的调节蛋白参与各种其他细胞和发育过程,并有助于清除细胞内病原体。自噬的生理作用和病理影响可以在从酵母到小鼠等不同的真核模型系统中进行有效研究。斑马鱼(Danio rerio)是一种仅在最近才应用于自噬研究的易于处理的动物模型,它也有助于分析更具体的生物学过程,如组织再生。在本章中,我们概述了用于监测该脊椎动物体内自噬结构和检测自噬反应的主要方法和工具。我们重点介绍了用于探索自噬过程新的细胞和发育作用的遗传(功能)方法。