Center for Neuropharmacology and Neuroscience, Albany Medical College, Albany, New York 12208, USA.
J Biol Chem. 2013 May 10;288(19):13676-94. doi: 10.1074/jbc.M112.420786. Epub 2013 Mar 26.
Missense mutations in AHI1 result in the neurodevelopmental ciliopathy called Joubert syndrome.
Mutations in AHI1 decrease cilia formation, alter its localization and stability, and change its binding to HAP1 and NPHP1.
Mutations in AHI1 affect ciliogenesis, AHI1 protein localization, and AHI1-protein interactions.
This study begins to describe how missense mutations in AHI1 can cause Joubert syndrome. Mutations in AHI1 cause Joubert syndrome (JBTS), a neurodevelopmental ciliopathy, characterized by midbrain-hindbrain malformations and motor/cognitive deficits. Here, we show that primary cilia (PC) formation is decreased in fibroblasts from individuals with JBTS and AHI1 mutations. Most missense mutations in AHI1, causing JBTS, occur in known protein domains, however, a common V443D mutation in AHI1 is found in a region with no known protein motifs. We show that cells transfected with AHI1-V443D, or a new JBTS-causing mutation, AHI1-R351L, have aberrant localization of AHI1 at the basal bodies of PC and at cell-cell junctions, likely through decreased binding of mutant AHI1 to NPHP1 (another JBTS-causing protein). The AHI1-V443D mutation causes decreased AHI1 stability because there is a 50% reduction in AHI1-V443D protein levels compared with wild type AHI1. Huntingtin-associated protein-1 (Hap1) is a regulatory protein that binds Ahi1, and Hap1 knock-out mice have been reported to have JBTS-like phenotypes, suggesting a role for Hap1 in ciliogenesis. Fibroblasts and neurons with Hap1 deficiency form PC with normal growth factor-induced ciliary signaling, indicating that the Hap1 JBTS phenotype is likely not through effects at PC. These results also suggest that the binding of Ahi1 and Hap1 may not be critical for ciliary function. However, we show that HAP1 has decreased binding to AHI1-V443D indicating that this altered binding could be responsible for the JBTS-like phenotype through an unknown pathway. Thus, these JBTS-associated missense mutations alter their subcellular distribution and protein interactions, compromising functions of AHI1 in cell polarity and cilium-mediated signaling, thereby contributing to JBTS.
AHI1 中的错义突变导致称为杰伯综合征的神经发育性纤毛病。
AHI1 中的突变会减少纤毛的形成，改变其定位和稳定性，并改变其与 HAP1 和 NPHP1 的结合。
AHI1 中的突变会影响纤毛发生，AHI1 蛋白的定位和 AHI1 蛋白相互作用。
这项研究开始描述 AHI1 中的错义突变如何导致杰伯综合征。 AHI1 中的突变导致杰伯综合征（JBTS），这是一种神经发育性纤毛病，其特征为中脑-后脑畸形和运动/认知缺陷。在这里，我们表明，来自患有 JBTS 和 AHI1 突变的个体的成纤维细胞中初级纤毛（PC）的形成减少。 AHI1 中导致 JBTS 的大多数错义突变发生在已知的蛋白结构域中，但是 AHI1 中的常见 V443D 突变发生在没有已知蛋白基序的区域中。我们表明，转染 AHI1-V443D 或新的导致 JBTS 的突变 AHI1-R351L 的细胞中，AHI1 在 PC 的基底体和细胞-细胞连接处的定位异常，这可能是由于突变 AHI1 与 NPHP1（另一种导致 JBTS 的蛋白）的结合减少所致。 AHI1-V443D 突变导致 AHI1 稳定性降低，因为与野生型 AHI1 相比，AHI1-V443D 蛋白水平降低了 50％。亨廷顿蛋白相关蛋白-1（Hap1）是一种调节蛋白，可与 Ahi1 结合，并且已经报道 Hap1 敲除小鼠具有类似 JBTS 的表型，这表明 Hap1 在纤毛发生中起作用。缺乏 Hap1 的成纤维细胞和神经元形成具有正常生长因子诱导的纤毛信号传导的 PC，表明 Hap1 的 JBTS 表型可能不是通过 PC 的作用。这些结果还表明，Ahi1 和 Hap1 的结合可能不是纤毛功能所必需的。但是，我们表明 HAP1 与 AHI1-V443D 的结合减少，这表明这种改变的结合可能通过未知途径导致类似 JBTS 的表型。因此，这些与 JBTS 相关的错义突变改变了它们的亚细胞分布和蛋白相互作用，从而损害了 AHI1 在细胞极性和纤毛介导的信号传导中的功能，从而导致 JBTS。
