mRNA as a mediator of immunological information transfer in vivo

描述（由申请人提供）：mRNA的体内注射触发对编码蛋白的免疫应答，并赋予针对疾病的保护。由于这个原因和其他原因，RNA疫苗具有巨大的临床潜力，因此我们了解它们如何诱导免疫是很重要的。我假设RNA疫苗可能利用了一种细胞类型和生物学途径，这种细胞类型和生物学途径已经进化到专门捕获、内化和翻译mRNA。我认为，这一途径有助于将免疫信息转移到未感染的树突状细胞（DC），从而在调节CD 8 + T细胞对许多病毒和细菌感染的反应中发挥关键作用。几乎所有的急性病毒感染诱导强烈的CD 8 + T细胞反应，这是启动时，na？ve CD 8 + T细胞通过与表达适当共刺激分子的DC表面上的MHC I类/表位肽复合物接触而活化。然而，许多病毒不感染DC;并且一些感染DC的病毒还编码相当有效地抑制MHC I类呈递的蛋白质。这些事实提出了一个难题：这些病毒编码的表位如何有效地被DC提呈？答案来自于交叉呈现的识别，如果它导致触发 初始CD 8 + T细胞引起交叉致敏。然而，两个体内观察结果表明，交叉呈递/交叉引发（下文称为CP）并不总是高效的。首先，肠道病毒复制到非常高的滴度，并诱导CD 4 + T细胞和抗体，但（独特的急性病毒感染），他们完全避免触发na？ve CD 8 + T细胞。其次，细胞外细菌感染，其中微生物蛋白质非常丰富，不会诱导强烈的CD 8 + T细胞反应。由于下面描述的原因，我假设这两个观察结果可以通过提出免疫信息向未感染的DC的一些转移可能依赖于mRNA（而不是蛋白质）来解释。因此，这项提议有两个目标：第一，评估裸RNA如何诱导免疫。第二，为了检验假设，在大多数微生物感染期间，mRNA被转移到未感染的DC;如果mRNA在其中被翻译，则编码的蛋白质 将到达I类MHC途径，诱导强烈的CD 8 + T细胞反应;我将这种机制命名为TATOR（RNA的转移和翻译）。相反，如果mRNA不能被翻译，则生物体不能被CD 8 + T细胞检测到。因此，其mRNA的特定特征使得肠道病毒和细胞外细菌对CD 8 + T细胞不可见。目标1.鉴定和表征参与RNA触发免疫的DC亚群。目标2.确定mRNA调控序列是否解释了为什么胞外细菌不能诱导强烈的CD 8 + T细胞应答。

DESCRIPTION (provided by applicant): In vivo injection of mRNA triggers immune responses to the encoded proteins, and confers protection against disease. For this and other reasons, outlined below, RNA vaccines have great clinical potential, so it is important that we understand how they induce immunity. I hypothesize that RNA vaccines may be exploiting a cell type, and biological pathway, that have evolved specifically to capture, internalize, and translate mRNA. I propose that this pathway facilitates the transfer of immunological information into uninfected dendritic cells (DCs), thereby playing a key part in regulating CD8+ T cell responses to many viral and bacterial infections. Almost all acute virus infections induce strong CD8+ T cell responses, which are initiated when na?ve CD8+ T cells are activated by contact with an MHC class I / epitope peptide complex on the surface of DCs that express appropriate costimulatory molecules. However, many viruses do not infect DCs; and some viruses that infect DCs also encode proteins that quite effectively inhibit MHC class I presentation. These facts posed a puzzle: how could epitopes encoded by these viruses be effectively presented by DCs? The answer came with the identification of cross-presentation which, if it results in the triggering of naive CD8+ T cells, causes cross-priming. However, two in vivo observations show that cross-presentation/cross-priming (hereinafter, CP) is not always highly-efficient. First, enteroviruses replicate to very high titers and induce CD4+ T cells and antibodies, yet (unique among acute virus infections) they completely avoid triggering na?ve CD8+ T cells. Second, extracellular bacterial infections, in which microbial protein is hugely abundant, do not induce strong CD8+ T cell responses. For reasons described below, I hypothesize that both observations can be explained by proposing that some transfer of immunological information into uninfected DCs may rely on mRNA (rather than protein). So, this proposal has two goals: First, to evaluate how naked RNA induces immunity. Second, to test the hypothesis that, during most microbial infections, mRNA is transferred to uninfected DCs; if it is translated therein, the encoded protein will reach the class I MHC pathway, inducing strong CD8+ T cell responses; I have named this mechanism TATOR (transfer and translation of RNA). Conversely, if the mRNA cannot be translated, the organism is undetectable by CD8+ T cells. Thus, the specific characteristics of their mRNAs renders enteroviruses and extracellular bacteria invisible to CD8+ T cells. Aim 1. To identify and characterize the DC subset that is involved in RNA-triggered immunity. Aim 2. To determine if mRNA regulatory sequences explain why extracellular bacteria fail to induce strong CD8+ T cell responses.