Agrobacterium tumefaciens is the causal agent of crown gall disease. The bacterium is capable of transferring a segment of single-stranded DNA (ssDNA) into recipient cells during the transformation process, and it has been widely used as a genetic modification tool for plants and nonplant organisms. Transferred DNA (T-DNA) has been proposed to be escorted by two virulence proteins, VirD2 and VirE2, as a nucleoprotein complex (T-complex) that targets the host nucleus. However, it is not clear how such a proposed large DNA–protein complex is delivered through the host nuclear pore in a natural setting. Here, we studied the natural nuclear import of the Agrobacterium-delivered ssDNA-binding protein VirE2 inside plant cells by using a split-GFP approach with a newly constructed T-DNA–free strain. Our results demonstrate that VirE2 is targeted into the host nucleus in a VirD2- and T-DNA–dependent manner. In contrast with VirD2 that binds to plant importin α for nuclear import, VirE2 directly interacts with the host nuclear pore complex component nucleoporin CG1 to facilitate its nuclear uptake and the transformation process. Our data suggest a cooperative nuclear import model in which T-DNA is guided to the host nuclear pore by VirD2 and passes through the pore with the assistance of interactions between VirE2 and host nucleoporin CG1. We hypothesize that this large linear nucleoprotein complex (T-complex) is targeted to the nucleus by a “head” guide from the VirD2–importin interaction and into the nucleus by a lateral assistance from the VirE2–nucleoporin interaction.

根癌农杆菌是冠gall病的病原体。该细菌能够在转化过程中将一段单链DNA（ssDNA）转移到受体细胞中，并且已被广泛用作植物和非植物生物的遗传修饰工具。已经提出，转移的DNA（T-DNA）将由两种毒力蛋白VirD2和VirE2陪伴，作为靶向宿主细胞核的核蛋白复合物（T-复合物）。但是，尚不清楚在自然环境中如何通过宿主核孔递送拟议的大型DNA-蛋白质复合物。在这里，我们研究了农杆菌的天然核输入通过使用分离的GFP方法和新构建的无T-DNA菌株，在植物细胞内提供了ssDNA结合蛋白VirE2。我们的结果表明，VirE2以依赖VirD2和T-DNA的方式靶向宿主核。与VirD2绑定到植物输入素α进行核输入相反，VirE2直接与宿主核孔复合物成分核孔蛋白CG1相互作用，以促进其核吸收和转化过程。我们的数据表明了一种合作的核输入模型，其中T-DNA被VirD2引导至宿主核孔，并在VirE2与宿主核孔蛋白CG1之间的相互作用的帮助下穿过孔。