Genetic dissection of Sle2 contribution to SLE pathogenesis

描述（由申请人提供）：本提案的目的是在NZM2410小鼠模型中确定负责Sle2狼疮相关表型的基因，并表征这些基因促进狼疮发病的机制。因此，Sle2，一个影响B细胞发育和功能的基因位点，在自身免疫发病机制中起主要作用。我们最近确定了三个独立的Sle2区域，Sle2a， Sle2b和Sle2c，它们影响B-1a细胞池的大小，但其中只有两个，Sle2a和Sle2b参与自身免疫发病机制。Sle2通过多种机制影响perC B-1a细胞的数量，这些机制可以通过三个Sle2位点中的每一个独立介导。最后，我们的长期合作伙伴Dr. Chandra Mohan （UTSW）利用56R抗dna重链转基因模型证明了Sle2介导了耐受性的破坏。基于这些最近的结果和我们一直用来表征Sle1基因的策略，我们提出了以下三个目标来识别Sle2基因：生成Sle2a、Sle2b和Sle2c位点的高分辨率遗传图谱。我们会生产B6。Sle2基因重组亚株，筛选增加的perC B-1a细胞池。这一过程将反复进行，直到每个位点被映射到< 0.5 cM的临界间隔。此外，我们将利用每个Sle2基因座与其他SLE易感基因座之间的相互作用作图方法，监测B-1a表型与狼疮发病机制的共分离。2. 表征与3个Sle2位点相关的表型。在进行基因定位工作的同时，我们将完善每个Sle2位点的表型定义。这一目的有两个目标，一是阐明这些基因座参与疾病机制的机制，二是促进aim 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细胞池的每种机制分配到特定的Sle2位点，2)比较表达每个Sle2位点的perC B-1a细胞的基因表达谱，以及3)使用56R模型确定三个Sle2位点中哪一个导致对核抗原的耐受性丧失。3. 鉴定Sle2a、Sle2b和Sle2c对应的基因。目标1和目标2将提供每个Sle2位点的位置和功能候选基因列表。B6与B6基因序列及表达多态性。Sle2同源菌株将对这些基因进行系统的鉴定。这些多态性相对于B细胞功能的功能将在同源菌株之间进行评估。功能多态性的存在与在临界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等位基因负责相应的SLE易感性表型。

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.