MHC characterization in Chinese rhesus macaques

描述(由申请人提供)：我们将通过测试艾滋病感染的中国恒河猴细胞表达感兴趣的特定等位基因来确定人类白细胞抗原类似物是否扩展到细胞反应性。我们还将确定在一组受感染动物中常见的MHC等位基因 并对它们进行表征，以识别SIV特异性表位。我们建议扩大和继续我们在以前的区域赠款中所做的工作。在最初的拨款中，我们基于MHC-肽结合基序的定义，以及来自体外MHC-肽结合分析的数据，对恒河猴进行了表征，并在人类和恒河猴中鉴定了一组类似的MHC分子。类似的MHC分子被定义为与高度物种间交叉反应(很大程度上重叠的肽结合谱系)相关的MHC分子。我们现在建议使用由下一代测序组成的新技术来推进这些分析，以识别一组感染SIV的中国恒河猴中的MHC分子。我们将通过测试表达感兴趣的特定等位基因的SIV感染的中国恒河猴的细胞来确定人类白细胞抗原类似物是否扩展到细胞反应性。此外，我们将识别这些感染动物中常见的MHC等位基因，并对它们进行特征分析，以确定SIV特异性表位。我们的目标也是从功能的角度理解人类白细胞抗原类似物，以及识别任何新的普遍表达的MHC I类等位基因。因此，我们提出了以下具体目标：1)利用下一代测序技术在一组感染SIVmac239的中国恒河猴中鉴定MHC I类等位基因的完整集合。2)研究中国恒河猴MHC：多肽结合基序及其相关的功能性免疫反应。华盛顿国家灵长类动物中心正在对56只中国恒河猴进行疫苗研究。我们已经有了关于这些动物子集的MHC初步数据，这是我们之前区域拨款的一部分。这些研究是使用传统的桑格测序方法进行的。我们现在将在[MiSeq平台]上使用下一代测序来扩展这些工作的MHC分型。在对这56只动物进行MHC鉴定后，我们将进行进一步的研究，以确定细胞免疫反应的特征。我们将收到PBMC，我们将用它来分析免疫反应。根据这些数据，我们将能够确定中国恒河猴MHC等位基因与HLA等位基因相似的背景下的细胞反应性，以及是否存在额外的高频等位基因。

DESCRIPTION (provided by applicant): We will determine if HLA analogy extends to cellular reactivity by testing cells from AIDS- infected Chinese rhesus macaques expressing specific alleles of interest. We will also identify MHC alleles that are common in a set of infected animals and characterize them to identify SIV-specific epitopes. We propose to expand and continue the work we performed in our previous AREA grant. In the original grant, we characterized rhesus macaques based on the definition of MHC-peptide binding motifs, data from in vitro MHC-peptide binding assays, and have identified sets of analogous MHC molecules in humans and rhesus macaques. Analogous MHC molecules are defined as MHC molecules associated with a high degree of interspecies cross-reactivity (largely overlapping peptide binding repertoires). We now propose to advance these analyses using new technology consisting of next generation sequencing, to identify MHC molecules in a set of SIV-infected Chinese rhesus macaques. We will determine if HLA analogy extends to cellular reactivity by testing cells from SIV- infected Chinese rhesus macaques expressing specific alleles of interest. Furthermore, we will identify MHC alleles that are common in these infected animals and characterize them to identify SIV- specific epitopes. Our goal is also to understand HLA analogy from a functional perspective as well as identify any new commonly expressed MHC class I alleles. Accordingly, we propose the following specific aims: 1) To identify the complete set of MHC class I alleles in a set of SIVmac239- infected Chinese rhesus macaques using next generation sequencing. 2) To characterize MHC:peptide binding motifs and associated functional immune responses specific in Chinese rhesus macaques. Fifty-six Chinese rhesus macaques are being studied as part of a vaccine study at Washington National Primate Center. We already have preliminary MHC data on a subset of these animals as part of our previous AREA grant. These studies were performed using traditional Sanger sequencing methods. We will now expand the MHC typing of these efforts using next generation sequencing on the [MiSeq platform]. Upon MHC identification of these 56 animals, we will perform additional studies to characterize cellular immune responses. We will receive PBMC which we will use to analyze immune responses. From these data, we will be able to determine cellular reactivity in the context of analogous MHC alleles in Chinese rhesus macaque to HLA alleles and whether there are additional high frequency alleles.