A glycine zipper motif is required for the translocation of a T6SS toxic effector into target cells

Abstract

Type VI secretion systems (T6SSs) can deliver diverse toxic effectors into eukaryotic and bacterial cells. Although much is known about the regulation and assembly of T6SS, the translocation mechanism of effectors into the periplasm and/or cytoplasm of target cells remains elusive. Here, we use the Agrobacterium tumefaciens DNase effector Tde1 to unravel the mechanism of translocation from attacker to prey. We demonstrate that Tde1 binds to its adaptor Tap1 through the N‐terminus, which harbors continuous copies of GxxxG motifs resembling the glycine zipper structure found in proteins involved in the membrane channel formation. Amino acid substitutions on G 39 xxxG 43 motif do not affect Tde1–Tap1 interaction and secretion but abolish its membrane permeability and translocation of its fluorescent fusion protein into prey cells. The data suggest that G 39 xxxG 43 governs the delivery of Tde1 into target cells by permeabilizing the cytoplasmic membrane. Considering the widespread presence of GxxxG motifs in bacterial effectors and pore‐forming toxins, we propose that glycine zipper‐mediated permeabilization is a conserved mechanism used by bacterial effectors for translocation across target cell membranes. image This study describes how the type VI secretion DNase effector Tde1 is translocated from the bacterial attacker into the competitor cell, demonstrating a new and maybe a conserved role of glycine zipper motif(s) in effector delivery. The N‐terminus of the Tde1 effector is necessary and sufficient for its loading onto the secretory machine and for secretion. A glycine zipper motif mediates cytoplasmic membrane permeabilization for target cell delivery. Glycine zipper‐mediated translocation may be a conserved membrane translocation mechanism for bacterial effectors.