Endogenous MHC class II antigen processing via autophagy

描述(申请人提供)：肿瘤细胞使用两个蛋白水解室来降解错误合成和其他受损的蛋白质、蛋白酶体和溶酶体。然后，当这些多肽分别通过MHC I类和II类呈现时，免疫系统会监测这两个系统的降解产物。Th1、CD4+T细胞识别MHC II类，CD8+T细胞识别MHC I类，可导致对感染细胞和恶性细胞的破坏。我们最近描述了致癌的Epstein Barr病毒(EBNA1)的核抗原1在自噬后进入MHC II类递呈。在这项应用中，我们现在建议从三个目的来研究自噬递送抗原是否构成内源性MHC II类抗原加工的一般途径。1.分析自噬水平及其与MHC-II类分子在人体组织中的重叠，在稳态状态下，以及在I型和II型干扰素免疫激活后，以及病毒感染中。我们将定量评估在有和没有免疫激活的不同人体组织中的自噬水平，并解决自噬小体和MHC II类负载囊泡之间的重叠问题。2.病毒和肿瘤抗原在自噬途径向MHC II类递呈后的特征。我们将调查其他病毒和肿瘤抗原是否遵循MHC II类抗原的自噬路线，其中CD4+T细胞经内源性处理后被识别。我们将构建EBNA1的缺失突变体，并分析它们在自噬后获得内源性MHC II类抗原处理的能力。这一分析将揭示针对该降解路径的底物的结构域。3.针对自噬降解和MHC II类递呈的内源性抗原。我们将针对已知的CD4+T细胞抗原到自噬小体，通过延长它们的半衰期来改善MHC II类递呈，因为长寿蛋白主要是通过自噬来处理的。此外，我们还将针对伴侣蛋白介导的信号肽自噬和利用LC3/Atg8蛋白的宏自噬，该蛋白在附着到自噬体内膜后在溶酶体中被部分降解。这些研究将使我们更好地了解免疫系统如何检测到肿瘤，以及我们如何增强对癌症的免疫反应。

DESCRIPTION (provided by applicant): Tumor cells use two proteolytic compartments to degrade incorrectly synthesized and other damaged proteins, proteasomes and lysosomes. The immune system then monitors the degradation products of these two systems when the peptides are presented via MHC class I and II, respectively. Recognition of MHC class II by Th1 CD4+ T cells and MHC class I by cytolytic CD8+ T cells will result in the destruction of infected and malignant cell. We have recently described that the nuclear antigen 1 of the oncogenic Epstein Barr virus (EBNA1) gains access to MHC class II presentation after autophagy. We now propose in this application to investigate in three aims if autophagic delivery of antigens constitutes a general pathway of endogenous MHC class II antigen processing. 1. Analysis of autophagy levels and overlap with MHC class II loading compartments in human tissues, during steady state and after immune activation by type I and II Interferons as well as viral infections. We will quantitatively assess the autophagy level in different human tissues with and without immune activation and address the overlap between autophagosomes and MHC class II loading vesicles. 2. Characteristics of viral and tumor antigens following an autophagic route to MHC class II presentation. We will investigate if other viral and tumor antigens, for which CD4+ T cell recognition after endogenous processing was demonstrated, follow the autophagic route for MHC class II presentation. We will construct deletion mutants of EBNA1 and analyze their capacity to access endogenous MHC class II antigen processing after autophagy. This analysis will reveal domains that target substrates for this degradation path. 3. Targeting of endogenous antigens for autophagic degradation and MHC class II presentation. We will target known CD4+ T cell antigens to autophagosomes for improved MHC class II presentation by extending their half-lifes, since long-lived proteins are primarily processed via autophagy. In addition, we will target for chaperone mediated autophagy with signal peptides and for macroautophagy with the LC3/Atg8 protein, which gets partially degraded in lysosomes after attachment to the inner autophagosome membrane. These studies will allow us to understand better how tumors can be detected by the immune system and how we can enhance immune responses to cancer.