Janardhan Ajit, Swigut Tomek, Hill Brian, Myers Michael P, Skowronski Jacek
Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, USA.
PLoS Biol. 2004 Jan;2(1):E6. doi: 10.1371/journal.pbio.0020006. Epub 2004 Jan 20.
The infectious cycle of primate lentiviruses is intimately linked to interactions between cells of the immune system. Nef, a potent virulence factor, alters cellular environments to increase lentiviral replication in the host, yet the mechanisms underlying these effects have remained elusive. Since Nef likely functions as an adaptor protein, we exploited a proteomic approach to directly identify molecules that Nef targets to subvert the signaling machinery in T cells. We purified to near homogeneity a major Nef-associated protein complex from T cells and identified by mass spectroscopy its subunits as DOCK2-ELMO1, a key activator of Rac in antigen- and chemokine-initiated signaling pathways, and Rac. We show that Nef activates Rac in T cell lines and in primary T cells following infection with HIV-1 in the absence of antigenic stimuli. Nef activates Rac by binding the DOCK2-ELMO1 complex, and this interaction is linked to the abilities of Nef to inhibit chemotaxis and promote T cell activation. Our data indicate that Nef targets a critical switch that regulates Rac GTPases downstream of chemokine- and antigen-initiated signaling pathways. This interaction enables Nef to influence multiple aspects of T cell function and thus provides an important mechanism by which Nef impacts pathogenesis by primate lentiviruses.
灵长类慢病毒的感染周期与免疫系统细胞之间的相互作用密切相关。Nef是一种强大的毒力因子，它会改变细胞环境以增加慢病毒在宿主体内的复制，但其作用机制仍不清楚。由于Nef可能作为一种衔接蛋白发挥作用，我们采用蛋白质组学方法直接鉴定Nef靶向的分子，以颠覆T细胞中的信号传导机制。我们从T细胞中纯化出一种主要的Nef相关蛋白复合物，使其纯度接近均一，并通过质谱鉴定其亚基为DOCK2-ELMO1（抗原和趋化因子启动的信号通路中Rac的关键激活剂）和Rac。我们发现，在没有抗原刺激的情况下，HIV-1感染后的T细胞系和原代T细胞中，Nef会激活Rac。Nef通过结合DOCK2-ELMO1复合物激活Rac，这种相互作用与Nef抑制趋化作用和促进T细胞活化的能力有关。我们的数据表明，Nef靶向一个关键开关，该开关在趋化因子和抗原启动的信号通路下游调节Rac GTP酶。这种相互作用使Nef能够影响T细胞功能的多个方面，从而提供了一种重要机制，通过该机制Nef影响灵长类慢病毒的发病机制。
