Genetic dissection of Sle2 contribution to SLE pathogenesis

描述(申请人提供)：这项建议的目的是确定在NZM2410小鼠模型中与Sle2狼疮相关的表型的相关基因，并表征这些基因促进狼疮发病的机制。因此，Sle2是一个影响B细胞发育和功能的遗传位点，在自身免疫发病机制中起着重要作用。我们最近发现了三个独立的Sle2区域，Sle2a，Sle2b和Sle2c，它们影响B-1a细胞池的大小，但只有两个区域，Sle2a和Sle2b与自身免疫发病有关。Sle2通过多种机制影响Perc B-1a细胞的数量，这些机制可能是通过三个Sle2基因座中的每一个独立地调节的。最后，我们的长期合作者钱德拉·莫汉博士(UTSW)已经证明，使用56R抗DNA重链转基因模型，Sle2介导了耐受性的破坏。基于这些最新的结果和我们一直使用的鉴定Sle1基因的策略，我们提出了以下三个目标来鉴定Sle2基因：1.构建Sle2a、Sle2b和Sle2c基因座的高分辨率遗传图谱。我们将产生B6.Sle2同源重组亚株，并对它们进行筛选，以增加Perc B-1a细胞库。此过程将反复进行，直到每个轨迹都映射到&lt；0.5 cm临界间隔。此外，我们将使用每个Sle2基因座与其他SLE易感基因座之间的相互作图方法来监测B-1a表型与狼疮发病机制的共同分离。2.研究3个Sle2基因座的表型特征。在遗传图谱工作的同时，我们将完善每个Sle2基因座的表型定义。这一目标具有双重目标，即阐明这些基因座在疾病机制中的作用机制，并促进目标3候选基因的选择。为了实现这一目标，我们将1)将Sle2增加B-1a细胞库的每一种机制分配给特定的Sle2基因座，2)比较表达每个Sle2基因座的perc B-1a细胞的基因表达谱，以及3)利用56R模型确定三个Sle2基因座中的哪一个导致对核抗原的耐受性丧失。3.确定与Sle2a、Sle2b和Sle2c对应的基因。AIMS 1和AIMS 2将提供每个Sle2基因座的位置和功能候选基因列表。B6和B6之间的序列和表达多态性将系统地表征这些基因的Sle2同源菌株。这些多态与B细胞功能相关的功能将在同源菌株之间进行评估。功能多态的存在和关键区间内的位置之间的一致性将提供强有力的证据，证明该特定基因的NZM2410等位基因与相应的SLE易感表型有关。

DESCRIPTION (provided by applicant): The purpose of this proposal is to identify the genes responsible for the Sle2 lupus-associated phenotypes in the NZM2410 murine model, and to characterize the mechanisms by which these genes contribute to lupus pathogenesis. Consequently, Sle2, a genetic locus that affects B cell development and function, plays a major role in autoimmune pathogenesis. We have recently identify three independent Sle2 regions, Sle2a, Sle2b, and Sle2c, that affect the size of the B-1a cell pool, but only two of them, Sle2a and Sle2b contribute to autoimmune pathogenesis. Sle2 affects the number of perC B-1a cells through multiple mechanisms, which could be mediated independently through each of the three Sle2 loci. Finally, our long-time collaborator Dr. Chandra Mohan (UTSW) has shown that Sle2 mediates a breach of tolerance using the 56R anti-DNA heavy chain transgenic model. Based on these recent results and on the strategy that we have been using to characterize the Sle1 genes, we propose the three following aims to identify the Sle2 genes: 1. To generate high resolution genetic maps of the Sle2a, Sle2b, and Sle2c loci. We will produce B6.Sle2 congenic recombinant sub-strains and screen them for increased perC B-1a cell pool. This process will be iterated until each locus has been mapped to a < 0.5 cM critical interval. In addition, we will monitor the co- segregation of the B-1a phenotype with the contribution to lupus pathogenesis using an interaction mapping approach between each of the Sle2 loci with other SLE susceptibility loci. 2. To characterize the phenotypes associated with each of the 3 Sle2 loci. Concurrent with the genetic mapping effort, we will refine the phenotypic definition of each Sle2 locus. This aim has the dual goal of elucidating the mechanisms by which these loci contribute to disease mechanisms, and facilitating the selection of candidate genes for Aim 3. To accomplish this goal, we will 1) assign each of the mechanisms by which Sle2 increases the B-1a cell pool to specific Sle2 loci, 2) compare the gene expression profile of perC B-1a cells expressing each Sle2 locus, and 3) determine which of the three Sle2 loci is responsible for the loss of tolerance to nuclear antigens using the 56R model. 3. To identify the genes corresponding to Sle2a, Sle2b, and Sle2c. Aims 1 and 2 will provide a list of positional and functional candidate genes for each of the Sle2 loci. Sequence and expression polymorphisms between the B6 and the B6.Sle2 congenic strains will be systematically characterized for these genes. The functionality of these polymorphisms relative to B cell functions will be then evaluated between the congenic strains. Congruity between the presence of a functional polymorphism and location within the critical interval will provide strong evidence that the NZM2410 allele of this specific gene is responsible for the corresponding SLE susceptibility phenotype.