Department of Anatomy, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan.
J Neurochem. 2011 Mar;116(6):1122-37. doi: 10.1111/j.1471-4159.2010.07167.x. Epub 2011 Jan 20.
SynArfGEF, also known as BRAG3 or IQSEC3, is a member of the brefeldin A-resistant Arf-GEF/IQSEC family and was originally identified by screening for mRNA species associated with the post-synaptic density fraction. In this study, we demonstrate that synArfGEF activates Arf6, using Arf pull down and transferrin incorporation assays. Immunohistochemical analysis reveals that synArfGEF is present in somata and dendrites as puncta in close association with inhibitory synapses, whereas immunoelectron microscopic analysis reveals that synArfGEF localizes preferentially at post-synaptic specializations of symmetric synapses. Using yeast two-hybrid and pull down assays, we show that synArfGEF is able to bind utrophin/dystrophin and S-SCAM/MAGI-2 scaffolding proteins that localize at inhibitory synapses. Double immunostaining reveals that synArfGEF co-localizes with dystrophin and S-SCAM in cultured hippocampal neurons and cerebellar cortex, respectively. Both β-dystroglycan and S-SCAM were immunoprecipitated from brain lysates using anti-synArfGEF IgG. Taken together, these findings suggest that synArfGEF functions as a novel regulator of Arf6 at inhibitory synapses and associates with the dystrophin-associated glycoprotein complex and S-SCAM.
SynArfGEF，也称为 BRAG3 或 IQSEC3，是布雷菲德菌素 A 抗性 Arf-GEF/IQSEC 家族的成员，最初是通过筛选与突触后密度部分相关的 mRNA 种类而鉴定的。在这项研究中，我们通过 Arf 下拉和转铁蛋白掺入测定证明了 synArfGEF 激活了 Arf6。免疫组织化学分析显示，synArfGEF 以与抑制性突触密切相关的点状存在于体和树突中，而免疫电镜分析显示，synArfGEF 优先定位于对称突触的突触后特化区。通过酵母双杂交和下拉测定，我们表明 synArfGEF 能够与位于抑制性突触的 utrophin/dystrophin 和 S-SCAM/MAGI-2 支架蛋白结合。双重免疫染色显示，synArfGEF 在培养的海马神经元和小脑皮质中分别与 dystrophin 和 S-SCAM 共定位。β-肌营养不良蛋白和 S-SCAM 都可以使用抗 synArfGEF IgG 从脑裂解物中免疫沉淀。总之，这些发现表明 synArfGEF 作为抑制性突触中 Arf6 的新型调节剂发挥作用，并与肌营养不良蛋白相关糖蛋白复合物和 S-SCAM 相关。
