Laminin γ3 chain binds to nidogen and is located in murine basement membranes
Laminin γ3 chain binds to nidogen and is located in murine basement membranes
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DOI：
10.1074/jbc.m501875200
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发表时间：
2005-06-10
影响因子：
4.8
通讯作者：
Miosge, N
中科院分区：
文献类型：
--
作者：
Gersdorff, N;Kohfeldt, E;Miosge, N
最近，在小鼠和人类中鉴定出一种新的层粘连蛋白γ 3链，并显示其具有与层粘连蛋白γ 1链相同的模块结构。我们在哺乳动物细胞中重组表达了γ 3链的两个片段。第一，结构域VI/V，由层粘连蛋白N-末端（结构域VI）和四个层粘连蛋白型表皮生长因子样（结构域V）和层粘连蛋白N-末端模块组成，被证明是层粘连蛋白自组装所必需的。另一个是结构域III 3 -5，它由三个层粘连蛋白型表皮生长因子样模块组成，并预测结合巢蛋白。γ 3 VI/V片段是层粘连蛋白-1聚合的不良抑制剂，γ 2 VI/ V片段也是如此。γ 3 III 3 -5片段以比γ 1 III 3 -5片段低的亲和力结合巢蛋白-1和巢蛋白-2。这些数据表明，含有γ 3链的层粘连蛋白可以独立于其他层粘连蛋白组装网络。针对γ 3 VI/V和γ 3 III 3 -5的多克隆抗体显示与层粘连蛋白的γ 1和γ 2链的同源片段没有交叉反应，并允许建立γ链特异性放射免疫测定和组织的光镜和电镜免疫染色。这表明在成年小鼠的各种组织提取物中，γ 3链的含量比γ 1链低20-100倍。γ 3链的表达具有高度的组织特异性。与早期的假设相反，针对γ 3链的抗体仅在成人和胚胎组织的基底膜区（如脑、肾、皮肤、肌肉和睾丸）显示光镜染色。超微结构免疫金染色定位γ 3链的基底膜，这些组织。
Recently a novel laminin gamma 3 chain was identified in mouse and human and shown to have the same modular structure as the laminin gamma 1 chain. We expressed two fragments of the gamma 3 chain in mammalian cells recombinantly. The first, domain VI/V, consisting of laminin N-terminal (domain VI) and four laminin-type epidermal growth factor-like (domain V) and laminin N-terminal modules, was shown to be essential for self-assembly of laminins. The other was domain III3-5, which consists of three laminin-type epidermal growth factor-like modules and is predicted to bind to nidogens. The gamma 3 VI/V fragment was a poor inhibitor for laminin-1 polymerization as was the gamma 2 VI/ V fragment. The gamma 3 III3-5 fragment bound to nidogen-1 and nidogen-2 with lower affinity than the gamma 1 III3-5 fragment. These data suggested that laminins containing the gamma 3 chain may assemble networks independent of other laminins. Polyclonal antibodies raised against gamma 3 VI/V and gamma 3 III3-5 showed no cross-reaction with homologous fragments from the gamma 1 and gamma 2 chains of laminin and allowed the establishment of gamma chain-specific radioimmunoassays and light and electron microscopic immunostaining of tissues. This demonstrated a 20-100-fold lower content of the gamma 3 chain compared with the gamma 1 chain in various tissue extracts of adult mice. The expression of gamma 3 chain was highly tissue-specific. In contrast to earlier assumptions, the antibodies against the gamma 3 chain showed light microscopic staining exclusively in basement membrane zones of adult and embryonic tissues, such as the brain, kidney, skin, muscle, and testis. Ultrastructural immunogold staining localized the gamma 3 chain to basement membranes of these tissues.