Department of Molecular Genetics, Kawasaki Medical School, 577 Matsushima, Kurashiki, Okayama 701-0192, Japan.
J Cell Sci. 2010 Mar 1;123(Pt 5):756-66. doi: 10.1242/jcs.060574.
Endosomal recycling of the mammalian iron transporter DMT1 is assumed to be important for efficient and rapid uptake of iron across the endosomal membrane in the transferrin cycle. Here, we show that the retromer, a complex that mediates retrograde transport of transmembrane cargoes from endosomes to the trans-Golgi network, is required for endosomal recycling of DMT1-II, an alternative splicing isoform of DMT1. Bacterially expressed Vps26-Vsp29-Vsp35 trimer, a retromer cargo recognition complex, specifically binds to the cytoplasmic tail domain of DMT1-II in vitro. In particular, this binding is dependent on a specific hydrophobic motif of DMT1-II, which is required for its endosomal recycling. DMT1-II colocalizes with the Vps35 subunit of the retromer in TfR-positive endosomes. Depletion of the retromer by siRNA against Vps35 leads to mis-sorting of DMT1-II to LAMP2-positive structures, and expression of siRNA-resistant Vps35 can rescue this effect. These findings demonstrate that the retromer recognizes the recycling signal of DMT1-II and ensures its proper endosomal recycling.
内体再循环哺乳动物铁转运蛋白 DMT1 被认为对于转铁蛋白循环中内体膜上铁的有效和快速摄取很重要。在这里，我们表明，参与跨膜货物从内体逆转运到反式高尔基体网络的复体型（retromer）对于 DMT1-II（DMT1 的一种选择性剪接同工型）的内体再循环是必需的。细菌表达的 Vps26-Vsp29-Vsp35 三聚体，是一种复体型货物识别复合物，在体外特异性结合 DMT1-II 的细胞质尾巴结构域。特别是，这种结合依赖于 DMT1-II 的特定疏水性基序，这是其内体再循环所必需的。DMT1-II 与内体转铁蛋白受体阳性的复体型的 Vps35 亚基共定位。通过针对 Vps35 的 siRNA 耗尽复体型会导致 DMT1-II 错误分拣到 LAMP2 阳性结构，并且表达 siRNA 抗性 Vps35 可以挽救这种效应。这些发现表明，复体型识别 DMT1-II 的再循环信号并确保其适当的内体再循环。
