Katoh Yohei, Shiba Yoko, Mitsuhashi Hiroko, Yanagida Yuko, Takatsu Hiroyuki, Nakayama Kazuhisa
Institute of Biological Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan.
J Biol Chem. 2004 Jun 4;279(23):24435-43. doi: 10.1074/jbc.M400059200. Epub 2004 Mar 26.
Tom1 (target of Myb1) is a protein of unknown function. Tom1 and its relative Tom1L1 have an N-terminal VHS (Vps27p/Hrs/Stam) domain followed by a GAT (GGA and Tom1) domain, both of which are also found in the GGA (Golgi-localizing, gamma-adaptin ear domain homology, ADP-ribosylation factor-binding protein) family of proteins. Although the VHS and GAT domains of GGA proteins bind to transmembrane cargo proteins and the small GTPase ADP-ribosylation factor, respectively, the VHS and GAT domains of Tom1 are unable to interact with these proteins. In this study, we show that the GAT domains of Tom1 and Tom1L1 interact with ubiquitin and Tollip (Toll-interacting protein). Ubiquitin bound the GAT domains of Tom1, Tom1L1, and GGA proteins, whereas Tollip interacted specifically with Tom1 and Tom1L1. Ubiquitin and Tollip bound to an overlapping region of the Tom1-GAT domain in a mutually exclusive manner. Tom1 was predominantly cytosolic when expressed in cells. On the other hand, Tollip was localized on early endosomes and recruited Tom1 and ubiquitinated proteins. These observations suggest that Tollip and Tom1 form a complex and regulate endosomal trafficking of ubiquitinated proteins.
Tom1（Myb1的靶标）是一种功能未知的蛋白质。Tom1及其相关蛋白Tom1L1具有一个N端VHS（Vps27p/Hrs/Stam）结构域，其后是一个GAT（GGA和Tom1）结构域，这两个结构域也存在于GGA（高尔基体定位、γ-衔接蛋白耳结构域同源性、ADP-核糖基化因子结合蛋白）家族蛋白中。尽管GGA蛋白的VHS和GAT结构域分别与跨膜货物蛋白和小GTP酶ADP-核糖基化因子结合，但Tom1的VHS和GAT结构域无法与这些蛋白相互作用。在本研究中，我们发现Tom1和Tom1L1的GAT结构域与泛素和Tollip（Toll相互作用蛋白）相互作用。泛素与Tom1、Tom1L1和GGA蛋白的GAT结构域结合，而Tollip则特异性地与Tom1和Tom1L1相互作用。泛素和Tollip以互斥的方式结合到Tom1-GAT结构域的重叠区域。在细胞中表达时，Tom1主要位于胞质溶胶中。另一方面，Tollip定位于早期内体，并募集Tom1和泛素化蛋白。这些观察结果表明，Tollip和Tom1形成复合物并调节泛素化蛋白的内体运输。
