肾病综合征(NS)是由肾小球滤过屏障功能障碍引起的常见肾脏疾病。在NS患者中发现的一些基因突变导致含肾脏锚蛋白重复序列(KANK)蛋白的氨基酸取代,它们是调节肌动蛋白聚合的支架蛋白,微管靶向,和细胞通过与各种分子结合而粘附,包括驱动蛋白运动蛋白KIF21A。然而,这些突变导致NS的机制尚不清楚.这里,我们意外地发现真核翻译起始因子4A1(eIF4A1)与NS相关的KANK2突变体(S684F)相互作用,但不与野生型蛋白相互作用。生化和结构分析显示,病理性突变诱导eIF4A1与KANK2在生理KIF21A结合位点的异常结合。竞争性结合测定进一步表明eIF4A1可以与KIF21A竞争以与KANK2的S684F突变体相互作用。在培养的小鼠足细胞中,该S684F突变体通过与eIF4A1结合而干扰了KANK2/KIF21A的相互作用,并且未能挽救已被KANK2敲除减少或在形态上改变的粘着斑或细胞粘附.这些结构性的,生物化学,和细胞结果不仅为S684F突变引起的足细胞缺陷提供了机械解释,但也显示了结合增益突变如何导致功能丧失效应。
Nephrotic syndrome (NS) is a common kidney disorder caused by dysfunction of the glomerular filtration barrier. Some genetic mutations identified in NS patients cause amino acid substitutions of kidney ankyrin repeat-containing (
KANK) proteins, which are scaffold proteins that regulate actin polymerization, microtubule targeting, and cell adhesion via binding to various molecules, including the kinesin motor protein KIF21A. However, the mechanisms by which these mutations lead to NS are unclear. Here, we unexpectedly found that the eukaryotic translation initiation factor 4A1 (eIF4A1) interacts with an NS-associated KANK2 mutant (S684F) but not the wild-type protein. Biochemical and structural analyses revealed that the pathological mutation induces abnormal binding of eIF4A1 to KANK2 at the physiological KIF21A-binding site. Competitive binding assays further indicated that eIF4A1 can compete with KIF21A to interact with the S684F mutant of KANK2. In cultured mouse podocytes, this S684F mutant interfered with the KANK2/KIF21A interaction by binding to eIF4A1, and failed to rescue the focal adhesion or cell adhesion that had been reduced or morphologically changed by KANK2 knockout. These structural, biochemical, and cellular results not only provide mechanistic explanations for the podocyte defects caused by the S684F mutation, but also show how a gain-of-binding mutation can lead to a loss-of-function effect.