Antigen processing and presentation of recombinant Listeria proteins

描述(申请人提供)：CD8+T淋巴细胞是主要的免疫效应细胞，对许多细胞内病原体，包括病毒，细菌和寄生虫，以及肿瘤做出反应。为了识别感染或肿瘤细胞，CD8+T细胞必须首先通过其T细胞受体与专业抗原提呈细胞(PAPC)表面显示的多肽-MHC(PMHC)复合体相互作用。这可以通过自然接触病原体或肿瘤抗原或故意将蛋白质引入pAPC的抗原处理机制来发生，就像在疫苗接种期间所做的那样。引入蛋白质以启动CD8+T细胞反应的最有效方法之一是使用活的、减毒的载体表达该蛋白质，例如胞内细菌单核细胞增生性李斯特菌。由于李斯特菌在最初感染后几分钟内进入宿主细胞胞浆，宿主细胞的抗原处理机制很容易获得细菌分泌的蛋白质，使李斯特菌成为激发CD8+T细胞反应的特别有效的载体。因此，我们一直在研究重组李斯特菌蛋白的加工机制，以呈现在MHC I类分子上。我们最近的研究发现，与内源性合成的蛋白质相比，重组李斯特菌抗原的加工和呈递方式有显着的不同。具体地说，我们发现，从分泌的李斯特菌蛋白中产生pMHC的速度是相同的，与其来源的蛋白质的细胞半衰期无关。我们的假设是，这些加工和呈现方面的差异反映了负责内源合成蛋白和外源细菌蛋白降解的细胞机制的真正差异。我们在本申请中提出的研究将集中于确定哪些因素导致加工过程中的这些差异。我们将通过1)确定蛋白质半衰期和加工效率的差异是否取决于细胞的内源蛋白质加工机制，以及2)确定李斯特菌在确定分泌重组蛋白质所遵循的加工途径中所起的作用。我们的研究应该会让我们更详细地了解I类抗原处理和呈递的机制。 公共卫生相关性：这些研究应该增加我们对以细胞内细菌病原体为载体表达的蛋白质的抗原处理和呈递机制的理解。蛋白质在受感染细胞中的降解和呈现方式会影响细胞免疫反应的质量和大小。这一知识对于我们理解来自病原体和恶性细胞的抗原蛋白是如何引起细胞免疫至关重要的。预计这些知识将有助于设计有效的疫苗。

DESCRIPTION (provided by applicant): CD8+ T lymphocytes are the primary immune effector cells that respond to many intracellular pathogens, including viruses, bacteria and parasites, as well as tumors. To recognize infected or neoplastic cells, CD8+ T cells must first be primed by interacting through their T cell receptor with peptide-MHC (pMHC) complexes displayed on the surface of professional antigen presenting cells (pAPC). This can occur either through natural exposure to the pathogen or tumor antigen or by intentionally introducing protein to the antigen processing machinery of a pAPC, as is done during vaccination. One of the most effective ways to introduce proteins for the purpose of priming a CD8+ T cell response is to express that protein using a live, attenuated vector such as the intracellular bacterium, Listeria monocytogenes. Because Listeria enters the host cell cytosol within minutes after initial infection, proteins secreted by the bacterium are readily accessible to the host cell's antigen processing machinery, making Listeria a particularly effective vector for eliciting CD8+ T cell responses. Therefore, we have been investigating the mechanisms involved in the processing of recombinant Listeria proteins for presentation on MHC class I molecules. Our recent studies have uncovered a striking difference in the way recombinant Listeria antigens are processed and presented when compared to endogenously synthesized proteins. Specifically, we have found that pMHC production from secreted Listeria proteins occurs at the same rate, independent of the cellular half-life of the protein from which it is derived. It is our hypothesis that these differences in processing and presentation reflect real differences in the cellular machinery responsible for the degradation of endogenously synthesized proteins vs. exogenously derived bacterial proteins. The studies we propose in this application will focus on determining what factors contribute these differences in processing. We will do this by 1) determining whether differences in protein half-life and processing efficiency are dependent on the endogenous protein processing machinery of the cell and 2) establishing the role Listeria localization plays in determining the processing pathway followed by secreted recombinant proteins. Our studies should provide us with a more detailed understanding of the mechanisms involved in class I antigen processing and presentation. PUBLIC HEALTH RELEVANCE: These studies should increase our understanding of the mechanisms of antigen processing and presentation of proteins expressed using an intracellular bacterial pathogen as a vector. How proteins are degraded and presented in infected cells can impact the quality and magnitude of the cellular immune response. This knowledge is critical to our understanding of how cellular immunity is elicited to antigenic proteins derived from pathogenic organisms and malignant cells. It is expected that this knowledge will aid in the design of effective vaccines.