Calkins Catharine C, Hoepner Bridgett L, Law Christine M, Novak Matthew R, Setzer Shannon V, Hatzfeld Mechthild, Kowalczyk Andrew P
Department of Dermatology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
J Biol Chem. 2003 Jan 17;278(3):1774-83. doi: 10.1074/jbc.M205693200. Epub 2002 Nov 7.
p0071, a member of the armadillo protein family, localizes to both adherens junctions and desmosomes in epithelial cells and exhibits homology to the adherens junction protein p120 and the desmosomal protein plakophilin-1. p0071 is also present at dermal microvascular endothelial intercellular junctions and colocalizes with VE-cadherin, an endothelium-specific cadherin that associates with both actin and intermediate filament networks. To define the role of p0071 in junction assembly, p0071 was tested for interactions with other components of the endothelial junctional complex. In transient expression assays, p0071 colocalized with and formed complexes with both VE-cadherin and desmoplakin. Deletion analysis using the yeast two-hybrid system revealed that the armadillo repeat domain of p0071 bound directly to VE-cadherin. Site-directed mutagenesis experiments demonstrated that p0071 and p120 bound to the same region on the cytoplasmic tail of VE-cadherin and that overexpression of p0071 could displace p120 from intercellular junctions. In contrast to VE-cadherin, desmoplakin was found to associate with the non-armadillo head domain of p0071. Cotransfections and triple-label immunofluorescence analysis revealed that VE-cadherin colocalization with desmoplakin in transfected COS cells required p0071, suggesting that p0071 may couple VE-cadherin to desmoplakin. Based on previous findings that both VE-cadherin and desmoplakin play central roles in vasculogenesis, these new results suggest that p0071 may play an important role in endothelial junction assembly and in the morphogenic events associated with vascular remodeling.
p0071是犰狳蛋白家族的成员，定位于上皮细胞的黏着连接和桥粒，与黏着连接蛋白p120和桥粒蛋白桥粒斑菲素蛋白-1具有同源性。p0071也存在于真皮微血管内皮细胞间连接中，并与VE-钙黏蛋白共定位，VE-钙黏蛋白是一种内皮细胞特异性钙黏蛋白，与肌动蛋白和中间丝网络都有关联。为了确定p0071在连接组装中的作用，检测了p0071与内皮连接复合体其他成分的相互作用。在瞬时表达试验中，p0071与VE-钙黏蛋白和桥粒斑蛋白共定位并形成复合物。使用酵母双杂交系统的缺失分析表明，p0071的犰狳重复结构域直接与VE-钙黏蛋白结合。定点诱变实验表明，p0071和p120与VE-钙黏蛋白细胞质尾部的同一区域结合，并且p0071的过表达可将p120从细胞间连接中置换出来。与VE-钙黏蛋白不同，发现桥粒斑蛋白与p0071的非犰狳头部结构域相关联。共转染和三标免疫荧光分析表明，在转染的COS细胞中，VE-钙黏蛋白与桥粒斑蛋白的共定位需要p0071，这表明p0071可能将VE-钙黏蛋白与桥粒斑蛋白连接起来。基于先前的发现，即VE-钙黏蛋白和桥粒斑蛋白在血管生成中都起核心作用，这些新结果表明，p0071可能在内皮连接组装以及与血管重塑相关的形态发生事件中起重要作用。
