A novel immunity system for bacterial nucleic acid degrading toxins and its recruitment in various eukaryotic and DNA viral systems

Zhang, Dapeng; Iyer, Lakshminarayan M.; Aravind, L.

Earlier characterized toxin systems such as the classical plasmid-encoded bacteriocins and the recently characterized CDI systems use thematically comparable, albeit biochemically distinct mechanisms for trafficking of nuclease toxins. While these systems have been used as models to understand bacterial protein trafficking, the complete set of events starting from the extrusion of the 'pro-toxin' by the producing cell to its recognition at the target cell surface and delivery into the target cell are only partially understood (8) . Classical plasmid-borne colicins and cognate bacteriocins from other bacteria do not have secretory mechanisms and their release appears to occur primarily through cell-lysis mediated by the colicin-release proteins (8) . Colicin-like bacteriocins are multidomain proteins with an extreme C-terminal toxin module, which is either a nuclease or a membraneperforating domain (e.g. colicin E1 and A) (8) . They typically possess two additional N-terminal modules, of which the first facilitates translocation across the target cell membrane and the second (i.e. the central module) facilitates binding to a membrane receptor on the target cell. These colicins hijack either the Tol or the Tondependent molecular import systems to enter the target cells (8) . The chromosomally encoded proteobacterial CDI system toxins do not require lysis; instead they are trafficked out of the cell which produces them via the twopartner-system that depends on the CdiB proteins belonging to the TpsB class of outer-membrane trafficking proteins (25) . These latter proteins contain N-terminal periplasmic polypeptide-transport-associated (POTRA) domains linked to a C-terminal b-barrel transmembrane domain. They recognize the secretory domains such as the TpsA-SD in the extreme N-terminal region of the CDI 'pro-toxins' to deliver them across the outer membrane of proteobacteria (64) . This N-terminal region is separated from the C-terminal regions by repetitive regions with RHS-or filamentous hemagglutinin-type repeats. Their uptake by the target cell is less-clearly understood. In the well-studied examples, the first step of this process appears to depend on the outer membrane-biogenesis protein BamA recognizing a conserved a-helical domain immediately N-terminal to the toxin module, with a VENN signature that overlaps with the PFAM model termed 'DUF638 0 . Subsequently the inner-membrane protein AcrB, a transporter, appears to be necessary for uptake into the target cell cytoplasm (25) . Additionally, it is posited that a proteolysis step at the cell surface releases just the C-terminal nuclease module for uptake by the target cell (25) . Thus, despite the differences between the CDI and classical colicin-like systems they share a common feature of the toxin activity being borne by the extreme C-terminal domain in a multidomain polypeptide. Further, the modules located immediately-N-terminal to the nuclease domain (e.g. the a-helical domain with the VENN motif $PFAM DUF638) are involved in association with receptors on the target cell. Hence, we term these domains collectively the pre-toxin (PT) domains. The extreme N-terminal domains appear to play a critical role in export from the host cell in the cases where lysis is not involved, i.e. typically chromosomally borne versions. These observations accordingly presented the organizational logic for these systems, wherein there are usually three functionally distinct sets of modules in the pro