Biological Mechanism of INF2-mediated FSGS

INF2介导的FSGS的生物学机制

• 批准号: 8318904
• 负责人: MARTIN R. POLLAK
• 金额: $ 41.1万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-15 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8318904
• 关键词: Actins; Alleles; Amino Acids; Binding; Biochemical; Biological; C-terminal; Cell physiology; Cells; Defect; Disease; Endoplasmic Reticulum; Engineering; Exons; Family; Family member; Focal Segmental Glomerulosclerosis; Genes; Glomerular Filtration Rate; Histologic; Human; Inherited; Injury; Kidney; Kidney Diseases; Kidney Failure; Lead; Mediating; Microfilaments; Microtubules; Modeling; Monomeric GTP-Binding Proteins; Mus; Mutate; Mutation; N-terminal; Pattern; Phenotype; Physiology; Point Mutation; Proteins; Proteinuria; RNA Splicing; Renal function; Role; Structure; Subgroup; Testing; Variant; base; depolymerization; formin-2; gain of function; glomerular function; human disease; in vivo; in vivo Model; loss of function; member; mouse model; mutant mouse model; podocyte; polymerization; public health relevance; research study; rho

DESCRIPTION (provided by applicant): We have identified a new gene which when mutated leads to a form of progressive kidney disease characterized by proteinuria, reduced glomerular filtration rate, and a histologic pattern of injury characterized by focal and segmental glomerulosclerosis (FSGS). This gene, INF2 (for Inverted Formin 2), is a member of the diaphanous subgroup of the formin family. INF2, like other members of the diaphanous formin subfamily, functions to accelerate actin filament polymerization. Unlike other members, INF2 can also accelerate actin depolymerization. Members of this subfamily of formins are able to autoinhibit their activity by an intramolecular interaction between two domains, the N-terminal DID (diaphanous inhibitory domain) and the C-terminal DAD (diaphanous autoregulatory domain). We have found point mutations in the DID of INF2 in eleven unrelated families with autosomal dominant FSGS. These mutations segregate with disease, are absent from controls, and alter highly conserved amino acids. The mode of inheritance, the absence of clear loss-of-function alleles, and the localization of all of the mutations to the DID, suggest a gain-of-function effect. In this application, we propose experiments to help define the mechanism by which INF2 mutations lead to human disease and to understand the role of INF2 in kidney function. In Aim 1, we propose to explore the effects of INF2 mutations on its biochemical functions. We will examine the hypothesis that FSGS- associated mutations in INF2 disrupt intramolecular interactions leading to functional changes in INF2. We will test the effects of FSGS-causing mutations on INF2 intramolecular interactions (DID with DAD), on the ability of INF2 to mediate actin filament polymerization and depolymerization, and the interactions of INF2 with actin filaments and microtubules. In Aim 2, we will define the effects of INF2 mutations on cellular functions. We will examine the hypothesis that INF2 mutations, by disinhibiting INF2 activity, alter actin- based cell function. We will study the role of INF2 in cells and the effect of INF2 mutations on actin-based structures, on actin dynamics, on the dynamics of the endoplasmic reticulum, and on interacting proteins. In Aim 3, we will develop and analyze two new INF2 mutant mouse models. We will use these models to test the role of INF2 in the podocyte and observe the effects of an INF2 point mutation in an in vivo model. We will engineer a mouse model with a disease-associated INF2 point mutation and also develop a podocyte-specific INF2 deficient mouse. We will examine the effect of INF2 mutation and deficiency on glomerular function in vivo. PUBLIC HEALTH RELEVANCE: We have identified a new focal segmental glomerulosclerosis (FSGS) gene. When mutated, this gene, INF2, causes kidney disease in humans. Better understanding how defects in this gene cause human disease will have significant and direct implications for understanding, and ultimately, treating, common forms of renal failure and renal failure progression.

描述（由申请人提供）：我们已经确定了一种新基因，当突变导致一种以蛋白尿为特征的进行性肾脏疾病形式，肾小球滤过率降低以及以局灶性和分段性肾小球乳化病（FSG）为特征的损伤的组织学模式。该基因INF2（用于倒数的formin 2）是formin家族的diaphanous子组的成员。像diaphanous formin亚家族的其他成员一样，INF2的功能可以加速肌动蛋白丝聚合。与其他成员不同，INF2还可以加速肌动蛋白解聚。 formins的该亚家族的成员能够通过两个结构域之间的分子内相互作用自动抑制其活性，N末端DID（透射抑制结构域）和C末端DAD（diaphanous daphanous自动调节结构域）。我们已经发现了十一个无关的FSG的无关家族中INF2的点突变。这些突变与疾病分离，对照不存在，并改变高度保守的氨基酸。遗传模式，缺乏明显的功能丧失等位基因以及所有突变对DID的定位表明了功能效果。在此应用中，我们提出了实验，以帮助定义INF2突变导致人类疾病并了解INF2在肾脏功能中的作用的机制。在AIM 1中，我们建议探索INF2突变对其生化功能的影响。我们将研究以下假设：INF2中FSGS相关的突变破坏了分子内相互作用，导致INF2功能变化。我们将测试引起FSG的突变对INF2分子内相互作用（与DAD一起使用），INF2介导肌动蛋白丝聚合和去聚合的能力的影响，以及INF2与肌动蛋白细丝和微管的相互作用。在AIM 2中，我们将定义INF2突变对细胞功能的影响。我们将研究以下假设：通过抑制INF2活性，INF2突变会改变基于肌动蛋白的细胞功能。我们将研究INF2在细胞中的作用以及INF2突变对基于肌动蛋白的结构，肌动蛋白动力学，对内质网的动力学以及对相互作用蛋白的影响。在AIM 3中，我们将开发和分析两个新的INF2突变小鼠模型。我们将使用这些模型来测试INF2在足细胞中的作用，并观察体内模型中INF2点突变的影响。我们将设计具有与疾病相关的INF2点突变的小鼠模型，并且还会发展出足细胞特异性INF2缺乏小鼠。我们将检查INF2突变和缺乏对体内肾小球功能的影响。 公共卫生相关性：我们已经确定了一种新的局灶性分段肾小球硬化（FSGS）基因。当突变时，该基因INF2在人类中引起肾脏疾病。更好地了解该基因中的缺陷如何导致人类疾病将对理解和最终处理肾衰竭和肾衰竭进展的常见形式具有重大和直接的影响。