SFB796 - Sub project C1
(sub project concluded on Sept 30 2010)


The structural and functional basis of reprogramming of vesicular transport by Salmonella enterica effector proteins


Project summary

Salmonella enterica is a facultative intracellular pathogen that resides and replicates within a membrane-bound compartment of host cells, the so-called "Salmonella-containing vacuole" or SCV. Intracellular Salmonella deploy a type III secretion system (T3SS) encoded by Salmonella Pathogenicity Island 2 (SPI2) to manipulate host cell functions and to achieve intracellular proliferation. Intracellular Salmonella induce a massive remodelling of the host cell endosomal system and the microtubule cytoskeleton. Furthermore, vesicular transport of infected cells is redirected towards the SCV, while antimicrobial factors are avoided. Of central importance for this pathogenic interference are the various effector proteins translocated by the SPI2 that act as toxins in the host cell. The proposed project is directed towards the understanding of the role of individual effector proteins, the identification of interacting host cell proteins and the structural basis of the function as effector proteins.

The work plan will comprise the following aspects:
1.   Understanding the role of SPI2 effector proteins in modification of cellular transport and remodelling of the endosomal system. Systematic analyses of alterations of the endosomal system and host cell transport will be performed using a live cell set-up. The detailed observation of changes to host cell functions should lead to refined models for the functions of individual effectors.
2.   Characterization of host cell functions involved in alteration of transport and remodelling of the endosomal system by Salmonella. Host cell functions that are predicted to be required for the manipulation by effector proteins of the SPI2-T3SS will be manipulated by microinjection of dominant-negative proteins or by a knock-down approach.
3.   Identification of host cell proteins interacting with SPI2 effector proteins. The host cell target proteins of SPI2-T3SS effector proteins are largely unknown and should be identified by combined genetic and protein interaction approaches. The screens will take into consideration the hydrophobic nature of the most important effector proteins.
4.   Structure/function relationship of SPI2 effector proteins. Effector proteins involved in the manipulation of the host cell cytoskeleton and vesicular transport are novel bacterial inventions without known functional domains or related proteins in other organisms. To achieve an understanding of the molecular function of these proteins, we will perform mutational analyses based on bioinformatics predictions for putative interaction domains. Improved bioinformatics will also be employed to define effector domains for recombinant expression and protein structure analyses. A future aim of the project is the determination of structures of effector/host cell target complexes.

The identification and analyses of effector/target interactions in the proposed project will lead to a molecular understanding of the intracellular strategies of an important intracellular pathogen.

Intracellular Salmonella enterica induce the remodeling of the host cell endosomal system. A transmission electron microscopy of an epithelial cell after infection with Salmonella enterica serovar Typhimurium wild type is shown. The bacterial cell is pseudo-colored green. Intracellular Salmonella reside a in specially membrane compartment termed 'Salmonella-containing vacuole'. Dependent on the function of effector proteins of the Salmonella Pathogenicity Island 2-encoded type III secretion system, intracellular Salmonella interfere with host cell vesicular transport and induce the formation of extensive tubular aggregations of endosomes, termed ‘Salmonella-induced filaments’ or SIF. The ultrastructure of a representative SIF is shown (pseudo-colored red). The image was modified from:  Rajashekar, R., Liebl, D., Seitz, A., and Hensel, M. (2008). Dynamic remodeling of the endosomal system during formation of Salmonella-induced filaments by intracellular Salmonella enterica. Traffic 9, 2100-2116.


Project relevant publications

  • Abrahams, G.L., Müller, P., and Hensel, M. (2006). Functional dissection of SseF, a type III effector protein involved in positioning the Salmonella-containing vacuole. Traffic 7, 950-965.
  • Kuhle, V., Abrahams, G.L., and Hensel, M. (2006). Intracellular Salmonella enterica redirect exocytic transport processes in a Salmonella pathogenicity island 2-dependent manner. Traffic 7, 716-730.
  • Kuhle, V., and Hensel, M. (2004). Cellular microbiology of intracellular Salmonella enterica: functions of the type III secretion system encoded by Salmonella pathogenicity island 2. Cell Mol Life Sci 61, 2812-2826.
  • Rajashekar, R., Liebl, D., Seitz, A., and Hensel, M. (2008). Dynamic remodeling of the endosomal system during formation of Salmonella-induced filaments by intracellular Salmonella enterica. Traffic 9, 2100-2116.
  • Xu, X., and Hensel, M. (2010). Systematic analysis of the SsrAB virulon of Salmonella enterica. Infect Immun 78, 49-58.