Gannage Monique, da Silva Rosa Barreira, Münz Christian
Viral Immunobiology, Institute of Experimental Immunology, University of Zürich, Zürich, Switzerland.
Methods Mol Biol. 2013;960:473-488. doi: 10.1007/978-1-62703-218-6_35.
Macroautophagy has recently emerged as an important catabolic process involved not only in innate immunity but also in adaptive immunity. Initially described to deliver intracellular antigens to MHC class II loading compartments, its molecular machinery has now also been described to enhance the delivery of extracellular antigens to MHC class II loading compartments by accelerating phagosome maturation. Therefore in pathological situations (viral or bacterial infections, tumorigenesis) the pathway might be involved in shaping CD4(+) T cell responses.In this chapter we describe three basic experiments for the monitoring and manipulation of macroautophagic antigen processing towards MHC class II presentation. Firstly, we will discuss how to monitor autophagic flux and autophagosome fusion with MHC class II loading compartments. Secondly, we will show how to target proteins to autophagosomes in order to monitor macroautophagy-dependent antigen processing via their enhanced presentation on MHC class II molecules to CD4(+) T cells. And finally, we will describe how macroautophagy can be silenced in antigen presenting cells, like human monocyte-derived dendritic cells (DCs).
巨自噬最近已成为一种重要的分解代谢过程，不仅参与固有免疫，还参与适应性免疫。最初它被描述为将细胞内抗原递送至MHC II类装载区室，现在其分子机制也被描述为通过加速吞噬体成熟来增强细胞外抗原向MHC II类装载区室的递送。因此，在病理情况下（病毒或细菌感染、肿瘤发生），该途径可能参与塑造CD4(+) T细胞反应。在本章中，我们描述了三个用于监测和操纵巨自噬性抗原加工以呈递至MHC II类分子的基本实验。首先，我们将讨论如何监测自噬流以及自噬体与MHC II类装载区室的融合。其次，我们将展示如何将蛋白质靶向自噬体，以便通过它们在MHC II类分子上增强呈递给CD4(+) T细胞来监测巨自噬依赖性抗原加工。最后，我们将描述如何在抗原呈递细胞（如人单核细胞衍生的树突状细胞（DC））中使巨自噬沉默。
