Defining the effector gene repertoire in systemic autoimmunity by ENU mutagenesis

描述（由申请人提供）：狼疮基本效应基因的鉴定为狼疮疾病过程中涉及的分子、机制和途径以及确定可能的治疗靶点提供了重要信息。然而，目前，这些基因的鉴定需要大量的育种，相对低效，并且受到敲除基因的可用性和基因功能已知要求的限制。最近，利用ENU诱变的正向遗传学方法已经成功地应用于大型和小型项目中，以解剖涉及正常和疾病状态的特定分子途径。虽然这一策略在SLE自发性模型中的应用受到许多因素的阻碍（包括交叉的要求，绘制ENU突变，必须具有相同的疾病严重程度），但已发表的和初步的数据为使用汞诱导的自身免疫模型（HgIA）来识别与自发性SLE相关的基本效应基因提供了强有力的支持。这种方法有可能识别HgIA发展所需的大多数主要基因，其中许多将适用于SLE，此外，与目前的方法相比，它不需要事先了解突变基因在自身免疫中的功能。通过潜在地开放整个基因组库进行筛选，这个拟议的项目可能会发现新的基因类别和新的机制。为了应用ENU方法来定义HgIA和SLE的基本效应基因，提出了两个具体目标。Aim 1将产生ENU突变体，能够抵抗氯化汞诱导的全身自身免疫。我们建议筛选总共800个enu突变配子，这将覆盖约12,700个隐性功能缺失突变，相当于大约一半的基因组。目标2将在选定的突变体中绘制和鉴定特定的enu诱导的遗传改变。拟议研究的结果将确定SLE中产生自身抗体所需的许多早期成分和机制，并可能确定新的基因和途径，这些基因和途径可能成为治疗干预的目标。

DESCRIPTION (provided by applicant): The identification of essential effector genes in lupus has provided important information both in terms of the molecules, mechanisms and pathways involved in the disease process and in identifying possible therapeutic targets. Currently, however, identification of such genes requires substantial breeding, is relatively inefficient, and is limited by both the availability of knockout genes and the requirement that the function of the gene be known. Recently, the forward genetics approach using ENU mutagenesis has been applied successfully in both large and small projects to dissect specific molecular pathways involved in normal and disease states. Although application of this strategy to spontaneous models of SLE is precluded by a number of factors (including the requirement that crosses, to map ENU mutations, must have homogeneous disease severity), published and preliminary data provide strong support for using the mercury-induced autoimmunity model (HgIA) to identify essential effector genes relevant to spontaneous SLE. This approach has the potential for identifying most of the major genes required for the development of HgIA, of which many will be applicable to SLE, and furthermore, in contrast to the current methods, it does not require prior knowledge about the function of the mutated genes in autoimmunity. By potentially opening up the entire genomic repertoire for screening, this proposed project is likely to identify new classes of genes and novel mechanisms. To apply the ENU approach to define essential effector genes in HgIA and SLE, two specific aims are proposed. Aim 1 will generate ENU mutants that are resistant to the systemic autoimmunity induced by mercuric chloride. We propose to screen a total of 800 ENU-mutated gametes, which would cover ~12,700 recessive loss-of-function mutations equivalent to about half the genome. Aim 2 will map and identify the specific ENU-induced genetic alteration in selected mutants. The results of the proposed studies should define many of the early components and mechanisms required for the production of autoantibodies in SLE and will likely identify novel genes and pathways that could be targeted for therapeutic intervention.