Fig. 1.

Genetic organization of vgrG1 and vgrG2 and their requirement in specific Tde toxin delivery. (A) Genetic organization of vgrG2 and hcp operons of A. tumefaciens strain C58. The genes are indicated by the annotated locus tag or designated name adapted from the literature (6, 37). The vgrG genes are in blue and the genes encoding the three toxins and their cognate immunity proteins are in red and green, respectively. (B) VgrG1 is required for secretion of Tde1 but not Tae. Western blot analysis of the total (T) and secreted (S) proteins from various A. tumefaciens strains expressing the plasmid control (p) or indicated plasmid. Protein names and molecular weight markers are indicated at the left and right, respectively. The soluble ActC protein was used as an internal nonsecreted protein control. (C and D) VgrG1 and VgrG2 are required for Tde1- and Tde2-dependent interbacterial competition activity, respectively. Various A. tumefaciens strains shown on the x axis were each mixed with ∆atu4349-tde1-tdi1 (C) or ∆tde2-tdi2 (D) at a 10:1 ratio and infiltrated into N. benthamiana leaves. The survival of ∆atu4349-tde1-tdi1 and ∆tde2-tdi2 collected at 0 and 24-h postinoculation (h) was quantified as cfu. Data are mean ± SEM (C, n = 4 biological repeats from two independent experiments; D, n = 5 biological repeats from three independent experiments). Significant difference compared with WT C58 at 24-h postinfiltration (*P ≤ 0.01). vgrG1 and vgrG2 are abbreviated as G1 and G2, respectively.