Abstract
The Type VI secretion system (T6SS) is a macromolecular machinery, widespread in Gram negative bacteria, which is used to deliver toxic proteins (“effectors”) directly into target eukaryotic or prokaryotic cells. As a result, this system plays a crucial role both in direct pathogenicity and in interbacterial interactions, including polymicrobial infections. T6SSs are encoded within large gene clusters containing at least thirteen conserved core components. These form a large trans-envelope machinery, including an expelled puncturing device consisting of a tube formed by hexameric rings of Hcp, topped by a VgrG trimer and further capped by a PAAR-repeat protein. Current models suggest that a variety of effectors can be injected in one single lethal shot, each either non-covalently associated or directly fused with one of the proteins forming the puncturing device. The opportunistic pathogen Serratia marcescens Db10 possesses a potent anti-bacterial T6SS. This T6SS is encoded by a gene large cluster in addition to several other loci encoding extra copies of some of the core components and multiple effectors.This study aimed to dissect the puncturing device of the T6SS of S. marcescens. The first part of this work was focused on determining the role of the three Hcp and two VgrG homologues in T6SS function and recruitment of different effectors. It was found that Hcp1 is essential to maintain T6SS–mediated antibacterial activity. Conversely, Hcp2 and Hcp3 seem to be dispensable for T6SS function. Next, it was found that S. marcescens can use either of the VgrG homologues to form a functional T6SS, albeit not with the same efficiency. To partially explain this, it was found that VgrG2 is essential for delivery of two PAAR-domain containing effectors, namely Rhs1 and Rhs2, and the novel effector Slp. In the second part of this study, the role of PAAR proteins in the T6SS was investigated. It was found that it is essential to have at least one PAAR protein to maintain T6SS function. Additionally, it was shown that S. marcescens can use different VgrG-PAAR combinations to form distinct variants of the puncturing device with differing anti-bacterial activities. Collectively, these results support the idea that bacteria can assemble a variety of functional T6SS puncturing devices, thereby increasing the range of toxins that can be delivered into a target cell and providing a large fitness advantage in a competitive environment.
| Date of Award | 2016 |
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| Original language | English |
| Awarding Institution |
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| Sponsors | Wellcome Trust |
| Supervisor | Sarah Coulthurst (Supervisor) |