Mouse modeling of a human STING gene variant for infectious disease

人类 STING 基因变体感染性疾病的小鼠模型

基本信息

批准号：
8282484
负责人：
John C Cambier
金额：
$ 19.26万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-01 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8282484
关键词：
Affect American Amino Acid Sequence Amino Acid Sequence Homology Amino Acids Animal Model Bacterial Infections C-terminal Communicable Diseases Cues Cyclic GMP Cytoplasmic Tail Defect Development Disadvantaged Disease Disease susceptibility Dominant-Negative Mutation Endoplasmic Reticulum Exons Fibroblasts Future Generations Genes Genetic Genetic Polymorphism Genetic Variation Goals Health Healthcare Host Defense Human Human Genetics Human Genome Immune response Immunity In Vitro Individual Infection Interferons Introns Knock-in Mouse Knowledge Listeria monocytogenes Mediating Membrane Methods Microbe Mitochondria Modeling Molecular Mus N-terminal Names Outer Mitochondrial Membrane Positioning Attribute Predisposition Production Proteins Relative (related person)Reporting Research Residual state Resistance to infection Risk Role Second Messenger Systems Signal Pathway Signal Transduction Site Sting Injury Structure Testing Time Transgenic Mice Transmembrane Domain Variant Viral Virus Diseases base experience gene function genetic variant genome wide association study human disease human study in vivo inhibitor/antagonist killings macrophage man microorganism mouse genome mouse model novel pathogen positional cloning predictive modeling response second messenger tool

项目摘要

DESCRIPTION (provided by applicant): Infectious diseases kill more people than any other single cause. Genome-Wide-Association-Studies in humans have demonstrated that genetic variations in genes involved in host defense can affect individuals' susceptibility to infection. I most cases, such reverse genetic approaches to understanding genetic contributions to disease are dependent on animal models that allow analysis of the effects of genetic variance in the absence of other variables. The lack of such models and the knowledge they yield poses a significant barrier for the future development of personalized treatment and health care for individuals carrying these genetic variations. Here we propose the construction of mouse gene knockin models to study the function of two variants of the human gene that encodes STING, a type I IFN stimulator that in mice is essential for defense against viral infections. We recently reported that there are two STING variants in man, each encoding three amino-acid changes (R71H-G230A-R293Q). We refer to these as WT and HAQ. Our in vitro studies have demonstrated that relative to WT the HAQ-STING variant has lost >90% of the ability to stimulate type I IFN production upon pathogen infection. Furthermore, the HAQ-STING variant has a dominant negative functional effect in vitro. Surprisingly, ~20% of Americans carry at least one copy of the HAQ-sting variant. It is our long-term goal to understand the molecular mechanisms and in vivo function of this frequent, potentially nonfunctional, human STING variant that may affect disease susceptibility of millions of Americans. To achieve this goal, we propose to develop two mouse models for the human STING variants. The two models will include 1) a minimalist knockin of SNPs into the mouse gene and 2) a knockin of SNPs plus unique human contextual sequence of potential importance in function. It is noteworthy that mouse and human STING proteins share 83% amino acid sequence homology. Thus in the first model (mHAQ mouse), we will make a STING knock-in in which the mouse equivalent amino acids will be changed to those in human HAQ-STING (Aim 1). In the second model we will accommodate the limited regions of non-homology, yet take advantage of the fact that mouse and human STING genes have the same exon - intron structure. We will make a STING knock-in that expresses a chimeric form of human-mouse STING, in which the entire N-terminal 172aa is replaced by the N-terminal of 173aa of human STING (cHAQ mouse) (Aim 2). This mouse will also contain G230A and R293Q. Appropriate WT mice will also be constructed. As time permits we will use these models to study the effects of HAQ on resistance to infection and on STING signaling function. PUBLIC HEALTH RELEVANCE: Sting is an important gene whose protein product functions in the control of our susceptibility to viral and bacterial infections. HAQ-STING is variant gene carried by ~20% of Americans. The protein encoded by HAQ- STING appears to be functionally defective, and therefore carriers of HAQ-STING may be at increased risk of infection. In this proposal, we will construct an animal model to definitively test the function of HAQ-STING encoded protein. In addition to advancing our understanding of the risk to human health posed by this gene variant,these animal models should be useful for developing and testing personalized treatments.

描述（由申请人提供）：传染病杀死的人比任何其他单一原因都多。人类全基因组的关联研究表明，宿主防御涉及的基因的遗传变异会影响个体对感染的敏感性。 i大多数情况下，这种反向理解对疾病的遗传贡献的反向遗传方法取决于动物模型，这些模型允许在没有其他变量的情况下分析遗传方差的影响。缺乏此类模型及其所产生的知识为携带这些遗传变异的个体的个性化治疗和医疗保健的未来发展带来了重大障碍。在这里，我们建议构建小鼠基因敲蛋白模型，以研究编码刺激的人类基因的两个变体的功能，该sting是一种I型IFN刺激剂，这对于防御病毒感染至关重要。我们最近报道说，人有两个刺激变体，每个变体都编码三个氨基酸变化（R71H-G230A-R293Q）。我们将其称为WT和HAQ。我们的体外研究表明，相对于WT，HAQ刺变变体损失了刺激病原体感染I型IFN产生的能力的90％。此外，HAQ-sting变体在体外具有主要的负功能效应。令人惊讶的是，约有20％的美国人至少携带了HAQ-sting变体的一份副本。我们的长期目标是了解这种频繁的，潜在的，可能会影响数百万美国人疾病易感性的频繁，可能无功能的人类刺痛变体的分子机制和体内功能。为了实现这一目标，我们建议为人刺变种开发两个鼠标模型。这两个模型将包括1）SNP的极简主义敲击素和2）SNP的敲击蛋白以及独特的人类情境在功能中的潜在重要性。值得注意的是，小鼠和人刺蛋白具有83％的氨基酸序列同源性。因此，在第一个模型（MHAQ小鼠）中，我们将进行刺痛，其中小鼠等效的氨基酸将变成人体HAQ刺的人（AIM 1）。在第二个模型中，我们将适应非体现的有限区域，但要利用小鼠和人刺基因具有相同的外显子内含子结构的事实。我们将进行一种表达人类鼠刺的嵌合体形式的刺痛，其中整个N末端172AA被173aa的人类刺（Chaq小鼠）的N末端取代（AIM 2）。该鼠标还将包含G230A和R293Q。还将构建适当的WT小鼠。在时间允许的情况下，我们将使用这些模型来研究HAQ对感染和刺激信号功能的影响。公共卫生相关性：Sting是一个重要的基因，其蛋白产物的作用在控制我们对病毒和细菌感染的敏感性方面。 Haq-sting是约20％的美国人携带的变异基因。通过HAQSTING编码的蛋白质似乎在功能上有缺陷，因此HAQ-Sting的载体可能会增加感染的风险。在此提案中，我们将构建一个动物模型，以确切地测试HAQ丁格编码蛋白的功能。除了促进我们对这种基因变体带来的人类健康风险的理解外，这些动物模型还应有助于开发和测试个性化治疗方法。