Broder Y C, Katz S, Aronheim A
Department of Molecular Genetics, Rappaport Family Institute for Research in the Medical Sciences, Faculty of Medicine, Technion-Israel Institute of Technology, P.O. Box 9649, Bat-Galim, Haifa, 31096, Israel.
Curr Biol. 1998 Oct 8;8(20):1121-4. doi: 10.1016/s0960-9822(98)70467-1.
The yeast two-hybrid system represents one of the most efficient approaches currently available for identifying and characterizing protein-protein interactions [1-4]. Although very powerful, this procedure exhibits several problems and inherent limitations [5]. A new system, the Sos recruitment system (SRS), was developed recently [6] based on a different readout from that of the two-hybrid system [6-8]. SRS overcomes several of the limitations of the two-hybrid system and thus serves as an attractive alternative for studying protein-protein interactions between known and novel proteins. Nevertheless, we encountered a number of problems using SRS and so have developed an improved protein recruitment system, designated the Ras recruitment system (RRS), based on the absolute requirement that Ras be localized to the plasma membrane for its function [9-10]. Ras membrane localization and activation can be achieved through interaction between two hybrid proteins. We have demonstrated the effectiveness of the novel RRS system using five different known protein-protein interactions and have identified two previously unknown protein-protein interactions through a library screening protocol. The first interaction (detailed here) is between JDP2, a member of the basic leucine zipper (bZIP) family, and C/EBPgamma, a member of the CCAAT/enhancer-binding protein (C/EBP) family. The second interaction is between the p21-activated protein kinase Pak65 and a small G protein (described in the accompanying paper by Aronheim et al. [11]). The RRS system significantly extends the usefulness of the previously described SRS system and overcomes several of its limitations.
酵母双杂交系统是目前用于鉴定和表征蛋白质-蛋白质相互作用的最有效方法之一[1-4]。尽管该方法非常强大，但仍存在一些问题和固有局限性[5]。最近开发了一种新系统，即Sos募集系统（SRS）[6]，其基于与双杂交系统不同的读出方式[6-8]。SRS克服了双杂交系统的一些局限性，因此是研究已知和新蛋白质之间蛋白质-蛋白质相互作用的有吸引力的替代方法。然而，我们在使用SRS时遇到了一些问题，因此基于Ras发挥功能绝对需要定位于质膜这一要求[9-10]，开发了一种改进的蛋白质募集系统，称为Ras募集系统（RRS）。Ras膜定位和激活可通过两种杂交蛋白之间的相互作用实现。我们使用五种不同的已知蛋白质-蛋白质相互作用证明了新型RRS系统的有效性，并通过文库筛选方案鉴定了两种以前未知的蛋白质-蛋白质相互作用。第一种相互作用（在此详细介绍）是碱性亮氨酸拉链（bZIP）家族成员JDP2与CCAAT/增强子结合蛋白（C/EBP）家族成员C/EBPγ之间的相互作用。第二种相互作用是p21激活的蛋白激酶Pak65与一种小G蛋白之间的相互作用（Aronheim等人在随附论文中描述[11]）。RRS系统显著扩展了先前描述的SRS系统的用途，并克服了其一些局限性。
