Kidney disease mechanisms associated with human genetic variation

慢性肾脏疾病(CKD)，像许多复杂的疾病表型一样，反映依赖的相互作用 遗传易感性和环境因素之间的关系。APOL1变异体解释了许多超额风险 非洲血统患者中的晚期非糖尿病慢性肾脏病。然而，只有部分具有风险基因的患者 患上肾脏疾病，表明遗传上位性或环境压力是触发变异的必要条件 APOL1依赖性肾损伤。为了研究APOL1变异是如何导致慢性肾脏病的，我们将重点放在HIV- 相关性肾病(HIVAN)，这种疾病与APOL1风险单倍型和 显然依赖于一个环境因素，即艾滋病毒-1感染。我们的初步数据显示APOL1是 在足细胞中表达，APOL1过表达激活自噬。尽管依赖于mTOR 抑制核心自噬可能导致糖尿病肾小球病变，这是一种与 APOL1变异体，我们提供的证据表明，APOL1与CKD的遗传关联源于APOL1- 依赖于对不同的、选择性的自噬途径的调节，从而导致病原体的破坏。 我们假设APOL1是一个自噬适配器，它系住了显示在 溶酶体与自噬蛋白LC3-II结合。APOL1的拴系功能是通过绑定一个 触发分子，HIV蛋白Nef，并导致携带 HIV病毒颗粒和蛋白质。变异的APOL1蛋白在功能上有缺陷，使HIV能够在 感染细胞并持续合成包括Nef在内的病毒蛋白。NEF阻止自噬消除 病毒，对HIVAN的发展至关重要。我们提出的实验解决了三个未回答的问题 关于APOL1在慢性肾脏病中的作用的新问题。APOL1是在肾脏中原位合成的吗？它是什么 亚蜂窝之家？循环或肾脏表达的APOL1变异体是否参与了肾脏疾病？会吗？ 对特定环境应激反应激活的自噬通路调节失调导致肾脏 APOL1风险基因携带者的疾病？这些问题是通过以下具体措施解决的 目的：1.检测APOL1在正常肾脏组织中的表达及细胞内定位，并确定APOL1在正常肾脏组织中的表达 风险基因和/或疾病诊断的定位不同；2.建立体内模型用于研究 APOL1与VAMP8的相互作用及其与HIV蛋白Nef的相互作用 检测正常和变异的APOL1对细胞系和培养的足细胞自噬的调节。这些 研究可以产生新的、基于机制的疗法，改善非裔美国人的健康差距 并确定人类慢性肾脏病的新机制。

Chronic kidney disease (CKD), like many complex disease phenotypes, reflects dependent interactions between genetic susceptibilities and environmental factors. APOL1 variants explain much of the excess risk of advanced non-diabetic CKD in patients of African ancestry. However, only some patients with risk genotypes develop kidney disease, suggesting that genetic epistasis or environmental stress is required to trigger variant APOL1-dependent kidney injury. To investigate how APOL1 variants result in CKD, we have focused on HIV- associated nephropathy (HIVAN), the disease most robustly associated with APOL1 risk haplotypes and clearly dependent on an environmental factor, HIV-1 infection. Our preliminary data demonstrate APOL1 is expressed in the podocyte, and APOL1 overexpression activates autophagy. Although mTor-dependent suppression of core autophagy may cause diabetic glomerulopathy, a renal pathology not associated with APOL1 variants, we provide evidence that the genetic association of APOL1 with CKD results from APOL1- dependent regulation of distinct, selective autophagy pathways, which result in the destruction of pathogens. We hypothesize that APOL1 is an autophagic adaptor that tethers a docking SNARE (VAMP8) displayed on lysosomes to the autophagosomal protein LC3-II. The tethering function of APOL1 is activated by binding a triggering molecule, the HIV protein Nef, and results in the selective degradation of autophagosomes carrying HIV viral particles and proteins. Variant APOL1 proteins are functionally defective, permitting HIV to persist in the infected cell and continue synthesis of viral proteins including Nef. Nef blocks autophagic elimination of the virus and is critical for development of HIVAN. Our proposed experiments address three unanswered and novel questions regarding APOL1 function in CKD. Is APOL1 synthesized in situ in kidney and what is its subcellular home? Does circulating or renal-expressed variant APOL1 mediate kidney disease? Does dysregulation of autophagy pathways activated in response to specific environmental stresses result in kidney disease in patients with APOL1 risk genotypes? These questions are addressed with the following Specific Aims: 1. Determine APOL1 expression and intracellular location in normal kidney, and determine if APOL1 localization varies with risk genotype and/or disease diagnosis; 2. Generate in vivo models for the study of APOL1 function; 3. Characterize protein interactions between APOL1 with VAMP8 and the HIV protein Nef and examine normal and variant APOL1 regulation of autophagy in cell lines and cultured podocytes. These studies can result in novel, mechanism-based therapies, improve health disparities in African American patients and identify novel mechanisms of human CKD.