Investigating the molecular mechanisms of prey recognition and prey killing in the soil bacterium Myxococcus xanthus

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The predatory soil bacterium Myxococcus xanthus kills other microorganisms to feed on their biomass. The multilayered predation strategy involves secretion of bactericidal antibiotics as well as bacteriolytic proteins, which accumulate within multicellular groups of M. xanthus and may enhance their predation efficiency. However, also single cells of M. xanthus are able to kill prey bacteria one-on-one in a contact-dependent manner. Here, a M. xanthus cell halts its motility when in contact with a prey cell and induces cell death and lysis within several minutes. Recently, we identified, by means of M. xanthus mutant analysis in different co-culture assays, two protein secretion systems that act together during cell contact-dependent killing of prey bacteria: a Tad-like system is required to induce prey cell death, while an atypical, needle-less type-3-secretion system (T3SS*) initiates prey cell disintegration. Mutation of both systems significantly reduces M. xanthus' ability to utilize biomass from bacteria and minimizes swarm expansion when growing on prey. Moreover, real time fluorescence microscopy revealed that components of both systems transiently accumulate at the predator-prey interface, presumably to deliver toxic effectors into the prey cell. To gain further insight into the functional interplay of the Tad-like and T3SS* systems during cell contact-dependent prey killing, the proposed research first addresses the translocation mechanism by the unusal, needle-less T3SS*. We aim to identify substrate proteins, to examine their translocation into the prey cell, and to determine the role of Tad components during this process. For this, we will take advantage on chaperone proteins, which likely recruit translocation substrates by direct protein-protein interaction (own preliminary data). Another focus of the project is the unknown mechanism of prey cell recognition. Our observation that the Tad/T3SS* killing machinery only accumulates upon contact with a prey cell, but not with another M. xanthus cell, provides a new read-out to investigate how M. xanthus indentifies a neighboring cell as prey. Here, we will systematically analyze Tad/T3SS* accumulation in M. xanthus paired with bacteria of different cell surface characteristics and of increasing phylogenetic distance. Furthermore, we will consider the putative role of known M. xanthus surface receptors, and compare predation to social incompatibility, which is mediated by the M. xanthus type-6-secretion system.