Bacteria live in complex communities and must compete with their neighbors for resources. They have therefore evolved multiple different competition systems in order to improve their fitness in dense, complex environments. The type VI secretion system (T6SS) is a Gram-negative weapon used to inhibit the growth of neighboring cells. It does this by delivering toxic effector proteins directly into neighbors using a speargun-like apparatus. A spear-shaped protein complex is propelled out of the T6SS-expressing cell, and this spear punctures a neighboring cell to deliver its toxic payload. Toxic effector proteins are bound either covalently or non-covalently to this secreted complex, and these toxins can target both periplasmic or cytoplasmic substrates, as well as membranes. Kin protect themselves from these effectors by expressing cognate immunity proteins to block effector activity.

This thesis explores several aspects of the T6SS in Enterobacter cloacae ATCC 13047. In Chapter 1, I provide a general introduction to the T6SS and covers topic such as the structure and assembly of the apparatus, regulation of T6SS loci and effectors, and effector diversity. I then describe the genetics of the T6SS in E. cloacae and explore what effectors are deployed in this system in Chapter 2. In Chapter 3, I focus on one particular effector deployed by this system, the type VI secretion system lipase effector, Tle, and investigate its intriguing reliance on its cognate immunity protein, Tli, for toxicity. Next, Chapter 4 discusses the role of rearrangement hotspotix (Rhs) proteins in the assembly of the T6SS apparatus. Chapter 5 then looks at the role of the Rhs accessory protein effector-associated gene with Rhs (EagR). Finally, I summarize my findings and discuss open questions in the field in Chapter 6.