The Department of Neuroscience, the Center for Hearing and Balance, The Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.
PLoS One. 2012;7(2):e31988. doi: 10.1371/journal.pone.0031988. Epub 2012 Feb 20.
Experiments utilizing the Looptail mutant mouse, which harbors a missense mutation in the vangl2 gene, have been essential for studies of planar polarity and linking the function of the core planar cell polarity proteins to other developmental signals. Originally described as having dominant phenotypic traits, the molecular interactions underlying the Looptail mutant phenotype are unclear because Vangl2 protein levels are significantly reduced or absent from mutant tissues. Here we introduce a vangl2 knockout mouse and directly compare the severity of the knockout and Looptail mutant phenotypes by intercrossing the two lines and assaying the planar polarity of inner ear hair cells. Overall the vangl2 knockout phenotype is milder than the phenotype of compound mutants carrying both the Looptail and vangl2 knockout alleles. In compound mutants a greater number of hair cells are affected and changes in the orientation of individual hair cells are greater when quantified. We further demonstrate in a heterologous cell system that the protein encoded by the Looptail mutation (Vangl2(S464N)) disrupts delivery of Vangl1 and Vangl2 proteins to the cell surface as a result of oligomer formation between Vangl1 and Vangl2(S464N), or Vangl2 and Vangl2(S464N), coupled to the intracellular retention of Vangl2(S464N). As a result, Vangl1 protein is missing from the apical cell surface of vestibular hair cells in Looptail mutants, but is retained at the apical cell surface of hair cells in vangl2 knockouts. Similarly the distribution of Prickle-like2, a putative Vangl2 interacting protein, is differentially affected in the two mutant lines. In summary, we provide evidence for a direct physical interaction between Vangl1 and Vangl2 through a combination of in vitro and in vivo approaches and propose that this interaction underlies the dominant phenotypic traits associated with the Looptail mutation.
利用携带vangl2 基因突变的 Looptail 突变鼠进行的实验对于平面极性的研究以及将核心平面细胞极性蛋白的功能与其他发育信号联系起来至关重要。最初被描述为具有显性表型特征，Looptail 突变表型的分子相互作用尚不清楚，因为突变组织中 Vangl2 蛋白水平显著降低或缺失。在这里，我们引入了一种 vangl2 敲除鼠，并通过杂交这两种品系并检测内耳毛细胞的平面极性，直接比较敲除和 Looptail 突变表型的严重程度。总体而言，vangl2 敲除表型比携带 Looptail 和 vangl2 敲除等位基因的复合突变体的表型更温和。在复合突变体中，受影响的毛细胞数量更多，并且当量化时单个毛细胞的取向变化更大。我们进一步在异源细胞系统中证明，Looptail 突变（Vangl2(S464N)）编码的蛋白通过 Vangl1 和 Vangl2(S464N)之间或 Vangl2 和 Vangl2(S464N)之间形成寡聚物，导致 Vangl1 和 Vangl2 蛋白向细胞表面的递送至细胞表面。与 Vangl2(S464N)的细胞内保留偶联，从而扰乱 Vangl1 和 Vangl2 蛋白的递送至细胞表面。因此，Vangl1 蛋白从 Looptail 突变体中的前庭毛细胞的顶端细胞表面缺失，但保留在 vangl2 敲除体中的毛细胞的顶端细胞表面。同样，Prickle-like2 的分布，一种假定的 Vangl2 相互作用蛋白，在两种突变体系中受到不同的影响。总之，我们通过体外和体内方法的组合提供了 Vangl1 和 Vangl2 之间直接物理相互作用的证据，并提出这种相互作用是 Looptail 突变相关显性表型特征的基础。
