Funtional analysis of Rhes, a novel Ras family protein, in neuronal cell

Rhes是Ras家族小GTP结合蛋白的成员。Rhes在脑中，特别是在纹状体中大量表达，因此被称为纹状体富集的Ras同源物。在本研究中，我们旨在确定Rhes的下游效应分子，从而深入了解Rhes的神经元功能。Rhes的效应子结合结构域不同于典型Ras同源物的那些，表明Rhes与非典型Ras效应子的相互作用。Ras家族中只有一个成员Rap 2与Rhes具有相似的效应子结合区。Rap 2与Rhes在效应子结合域的第8位上共享Phe残基，其中Ras具有Ser。Rap 2在脑中表达，并参与神经功能。因此，我们推断Rhes和Rap 2可以共享共同的非典型Ras效应子。我们使用酵母双杂交筛选和亲和纯化-质谱的方法来寻找Rlies或Rap 2结合蛋白。出乎意料的是，我们分离了Rhes和Rap 2的不同结合蛋白，如ARMS（用于Rhes）和MAP 4K 4（用于Rap 2）。ARMS是NGF受体的下游分子，而MAP 4K 4的果蝇同源物调节神经元轴突靶向。进一步的研究是必要的，以澄清这些发现的生理意义。

Rhes is a member of the Ras family small GTP-binding proteins. Rhes is expressed abundantly in brain, especially in striatum, and thus designated Ras homologue enriched in striatum. In the present study, we aimed at identifying a downstream effector molecule of Rhes thereby gaining insights into Rhes's neuronal function. Rhes's effector-binding domain is different from those of typical Ras homologue, suggesting interaction of Rhes with non-typical Ras effectors. Only one member of Ras family, Rap2, shares similar effector-binding region with Rhes. Rap2 shares Phe residue at the 8th position of effector-binding domain with Rhes, where Ras has Ser. Rap2 is expressed in brain and implicated in neuronal functions as well. We therefore reasoned that Rhes and Rap2 could share common non-typical Ras effectors. We used the yeast two-hybrid screening and affinity purification-mass spectrometry approach to search for Rlies-or Rap2-binding proteins. Unexpectedly, we isolated different binding proteins for Rhes and Rap2, such as ARMS(for Rhes) and MAP4K4(for Rap2). ARMS is a downstream molecule of NGF receptor, while MAP4K4's Drosophila homologue regulates neuronal axon targeting. Further studies are necessary to clarify physiological significances of these findings.