Elucidation of the role of 2 novel cross presentation genes

 描述（由申请人提供）：树突状细胞（DC）和巨噬细胞能够通过称为交叉呈递的过程从外部抗原获得并随后展示肽。这一过程是免疫系统识别并动员CD 8 T细胞对癌症、许多病毒感染和细胞内病原体做出反应的关键机制。因此，该途径对于免疫监视是重要的，并且是使疫苗能够引发CD 8 T细胞免疫的有吸引力的靶标。鉴于此，重要的是要阐明使发展中国家能够履行这一关键职能的机制，这是这项赠款的总体目标。为此，我们建议表征和阐明两个关键的和以前未知的新的交叉呈递基因的功能，我们发现在一个公正的，全基因组正向遗传筛选。我们的第一个目标是阐明Rab 39 A在交叉呈递中的作用。非常有趣的是，Rab 39 A是交叉呈递所必需的，而不是常规的MHC I或MHC II呈递。Rab蛋白是控制细胞中囊泡的形成、含量、运输和最终融合并由此决定内吞区室的组成的GTP酶。我们支持的假设是Rab 39 A通过运输XPT途径的基本组分来帮助在DC中形成专门的XPT区室。此外，考虑到它们在确定囊泡组成中的作用，Rab及其募集的效应物用作鉴定和分离独特的内吞隔室（例如早期内体、晚期内体、溶酶体、黑素体等）的标记物。在这种情况下，非常有趣的是，Rab 39 A仅存在于DC中的吞噬体的子集中。鉴于此，我们还假设Rab 39 A将标记XPT隔室。这些假设的重要性在于Rab 39 A可以解决MHC I和/或其他来自ER的组分如何进入吞噬体的谜团;澄清XPT在哪里进行及其与其他途径如MHC II的关系;和/或允许Rab 39 A+隔室的表征以识别途径中的新组分。本研究旨在验证这些假设，并更广泛地探索和阐明Rab 39 A在XPT中的作用。我们的第二个目标是阐明Loc 547349（Loc）在交叉呈递中的作用。我们的初步数据表明，这种分子是所需的交叉介绍，但不是经典的MHC I或MHC II抗原呈递。非常有趣的是，Loc是一种MHC编码的MHC I样分子，功能未知。这一发现非常有趣，因为许多MHC I和MHC II抗原呈递的关键基因都编码在MHC基因座中。此外，还有一种类似的分子HLA-DM，在MHC II途径中作为肽编辑器发挥作用，迄今为止在MHC I途径中还没有明确的对应物，但理论上需要这种交叉呈递活性。我们倾向于认为Loc在交叉呈递中的功能是帮助MHC I分子进入内吞区室，稳定它们，和/或用肽装载它们。本研究的目的是验证这一假设以及其他潜在的功能，并阐明位置在交叉呈递中的作用。

 DESCRIPTION (provided by applicant): Dendritic cells (DCs) and macrophages are capable of acquiring and then displaying peptides from external antigens through a process called cross presentation. This process is the key mechanism that allows the immune system to recognize and then mobilize a CD8 T cell response to cancers, many viral infections and intracellular pathogens. Consequently, this pathway is important for immune surveillance and is an attractive target to enable vaccines to elicit CD8 T cell immunity. Given this, it is important to elucidate te mechanisms that allow DCs to carry out this critical function and this is the overall goal of this grant. To this end, we propose to characterize and elucidate the function of two key and previously unsuspected novel cross presentation genes that we discovered in an unbiased, genome-wide forward genetic screen. Our first Aim will elucidate the role of Rab39A in cross presentation. Very interestingly, while Rab39A is required for cross presentation but not conventional MHC I or MHC II presentation. Rab proteins are GTPases that control the formation, content, trafficking and ultimate fusion of vesicles in cells and thereby determine the composition of endocytic compartments. Our favored hypothesis is that Rab39A is functioning to help form a specialized XPT compartment in DCs by trafficking essential components of the XPT pathway. Furthermore, given their role in determining the composition of vesicles, Rabs and their recruited effectors serve as markers to identify and isolate unique endocytic compartments (e.g. early endosomes, late endosomes, lysosomes, melanosomes, etc.). In this context, and very interestingly, Rab39A is only present in a subset of phagosomes in DCs. Given this, we also hypothesize that Rab39A will mark the XPT compartment. The importance of these hypotheses is that Rab39A could solve the mystery as to how MHC I and/or other components from the ER get to phagosomes; clarify where XPT is carried out and its relationship to other pathways such as MHC II; and/or allow the characterization of the Rab39A+ compartment to identify novel components in the pathway. This Aim will test these hypotheses and more generally explore and elucidate the role of Rab39A in XPT. Our second Aim will elucidate the role of Loc547349 (Loc) in cross presentation. Our preliminary data show that this molecule is required for cross presentation but not classical MHC I or MHC II antigen presentation. Very interestingly, Loc is an MHC-encoded MHC I-like molecule of unknown function. This discovery is very intriguing because many of the key genes for MHC I and MHC II antigen presentation are encoded in the MHC locus. Moreover, there is a similar molecule, HLA-DM, that operates as a peptide editor in the MHC II pathway and for which up until now has not had a clear counterpart in the MHC I pathway, yet there is a theoretical need for such an activity for cross presentation. Our favored hypotheses are that Loc functions in cross presentation to help traffic MHC I molecules to endocytic compartments, stabilize them, and/or load them with peptides. The goal of this aim is to test this hypothesis as well as other potential functions and elucidate the role of Loc in cross presentation.