An Rhs effector uses distinct target cell functions to intoxicate bacterial and fungal competitors

Many bacteria use Type VI secretion systems (T6SSs) to deliver toxic effector proteins into neighbouring bacterial or fungal cells as a means of inter-microbial competition. Compared with numerous antibacterial effectors, few antifungal effectors have been described. Furthermore, how T6SS-delivered effectors reach their site of action in different types of target cell remains poorly understood. Here, we combine structural biology with in vivo approaches to show that Rhs2 from Serratia marcescens Db10 is a dual-kingdom T6SS-dependent DNase effector which hijacks distinct, essential target cell functions in order to reach its site of action in bacterial and fungal cells. In bacterial cells, the Rhs2 toxin domain (Rhs2CT) interacts specifically with the elongation factor, EF-Tu, and, in sibling cells, interacts with the cognate immunity protein in an unusual manner. Interaction with EF-Tu is essential for T6SS-mediated intoxication of bacterial cells by Rhs2, but not for DNase activity or intoxication of fungal cells, implying it facilitates entry of Rhs2CT across the inner membrane to the cytoplasm. Alternatively, in fungal cells, Rhs2CT translocates to the nucleus using the nuclear import machinery. Our findings reveal how a single effector domain can act against targets with distinct cellular architectures and suggest that dual-kingdom effectors may occur widely.