News AG Bacterial Physiology

September 14, 2020
Structure and Function of Stator Units of the Bacterial Flagellar Motor

In a great collaboration with the group of Nicholas Taylor, we present the structure of nature’s smallest rotary motor - the stator units that power bidirectional rotation of the bacterial flagellum. Published in Cell!

The stator units (formed by the MotA–MotB membrane protein complex) use energy derived from the proton gradient across the inner membrane to power rotation of the flagellum. However, despite decades of research, their structure and molecular mode of action has remained a mystery. In this paper, we found that the stator unit consists of a dimer of MotB surrounded by a pentamer of MotA, which interacts with the rotor of the flagellum. Quite surprisingly, the stator units function as miniature rotary motors themselves, where a pentamer of MotA rotates around a MotB dimer.

See below for a movie of the mechanistic model for how the stator units can power bidirectional rotation of the flagellar motor.

April 24, 2020
Methylation of Salmonella Typhimurium flagella promotes bacterial adhesion and host cell invasion

We show that posttranslational methylation of the flagellar filament contributes to efficient adhesion of Salmonella Typhimurium to hydrophobic host cell surfaces. Published in Nature Communications!

It is known since 1959 that flagella of Salmonella Typhimurium are ɛ-N-methylated at lysine residues of flagellin via the methylase FliB. In this paper, we used mass-spectrometry and x-ray crystallography in collaboration with Charlotte Uetrecht and Michael Kolbe to determine the methylated lysine residues of both flagellins of Salmonella, FliC and FljB. We find that the flagellins are primarily methylated at surface-exposed residues. A Salmonella Typhimurium mutant of FliB, which is deficient in flagellin methylation, was outcompeted for gut colonization in a gastroenteritis mouse model. In support, the presence of methylated flagella facilitated bacterial invasion of epithelial cells. Finally, we found with the help of Yannick Rossez that lysine methylation increases the surface hydrophobicity of flagellin, and enhances flagella-dependent adhesion of Salmonella to phosphatidylcholine vesicles and epithelial cells. We therefore conclude that posttranslational methylation of flagellin promotes adhesion of Salmonella Typhimurium to hydrophobic host cell surfaces, and contributes to efficient gut colonization and host infection.

December 18, 2019
ERC consolidator grant

Very happy and honored to receive funding for our project BacNanoMachine from the European Research Council!

In the project BacNanoMachine, we aim to obtain a holistic understanding of the underlying principles that allow bacteria to control and coordinate the simultaneous self- assembly processes of several multi-component nanomachines within a single cell.
Despite being unicellular organisms of relatively small size, bacteria produce sophisticated nanomachines with a high degree of self-organization. The motility organelle of bacteria, the flagellum, is a prime example of complex bacterial nanomachines. Flagella are by far the most prominent extracellular structures known in bacteria and made through self-assembly of several dozen different kinds of proteins and thus represents an ideal model system to study sub-cellular compartmentalization and self-organization.
The flagellum can function as a macromolecular motility machine only if its many building blocks assemble in a coordinated manner. However, previous studies have focused on phenotypic and genetic analyses, or the characterization of isolated sub-components. Crucially, how bacteria orchestrate the many different cellular processes in time and space in order to construct a functional motility organelle remains enigmatic.
In the project BacNanoMachine, we will combine for the first time the visualization of the dynamic self-assembly of individual flagella with quantitative single-cell gene expression analyses, re-engineering of the genetic network and biophysical modeling in order to develop a biophysical model of flagella self-assembly.

May 14, 2019
New stimulated emission depletion (STED) microscope

We are happy to get support from the DFG to purchase a new superresolution microscope!
Superresolution microscopy is needed in order to investigate cellular structures and dynamic cellular processes below the Abbe diffraction limit. The new STED microscope will serve the need for superresolution light microscopy on a nanometer-scale (nanoscopy) of several research groups at the Institute for Biology.
Check out some example images below, where we have detected the flagellar filament and hook structure using STED microscopy and compared the images to the normal confocal microscopy mode. In contrast to the diffraction-limited confocal mode, STED microscopy allowed us to precisely measure on a nanometer-scale the size of the flagellar hook structure, which – according to electron microscopy experiments – grew to a length of ∼55 nm and substantially increased the spatial resolution of the immunostained flagellar filaments

Left: Unprocessed (raw) confocal image of co-stained flagellar hook- filament structures. Right: Unprocessed (raw) STED image of the same field of view.

September 6, 2018
Hook length of the bacterial flagellum is optimized for maximal motility performance

We demonstrate that bacteria control the optimal length of the flagellar hook structure to nanometer-scale in order to maximize stability of the flagellar bundle. Published in PLoS Biology in collaboration with Yann Dufour and Wilson Poon!

Many bacteria use flagella for directed movement in liquid environments. The flexible hook connects the membrane-embedded basal body of the flagellum to the long, external filament. Flagellar function relies on self-assembly processes that define or self-limit the lengths of major parts. The length of the hook is precisely controlled on a nanometer scale by a molecular ruler mechanism. However, the physiological benefit of tight hook-length control remained unclear. In this study, we show that the molecular ruler mechanism evolved to control the optimal length of the flagellar hook, which is consistent with efficient motility performance. These results highlight the evolutionary forces that enable flagellated bacteria to optimize their fitness in diverse environments and might have important implications for the design of swimming microrobots.

May 1, 2018
Regulation of flagella synthesis in response to cell envelope stress

We show that the degenerate EAL domain protein RflP responds to cell envelope stress and subsequently downregulates flagella biosynthesis. Published in mBio!

Salmonella enterica uses flagellum-mediated motility to reach preferred sites of infection. However, the flagellum also constitutes a prime target for the host’s immune response. Accordingly, the pathogen needs to determine the correct spatiotemporal stage of infection and control flagellar biosynthesis in a robust manner. We found that Salmonella responds to signals from cell envelope stress-sensing systems to turn off production of flagella. The degenerate EAL domain protein RflP is expressed in response to cell envelope stress sensed via the Rcs/RpoE pathways. RflP subsequently targets the flagellar master regulator FlhDC for proteolytic degradation. We speculate that downregulation of flagellum synthesis after cell envelope damage in hostile environments aids survival of Salmonella during late stages of infection and provides a means to escape recognition by the immune system.

November 28, 2017
Protein-conducting pore of the flagellum

In collaboration with Kelly T. Hughes and David F. Blair at the University of Utah, we show that the secretion pore of the flagellar type-III protein export apparatus is formed by the protein FliP. Published in Molecular Microbiology!

The flagellar type-III secretion system (T3SS) exports most extra-cytoplasmic components of the flagellum with remarkable speed and specificity. The membrane-embedded components of the T3SS are formed by FlhA, FlhB, FliP, FliQ and FliR. Here, we showed that the protein-conducting conduit is formed primarily, and possibly entirely, by FliP. We propose that a methionine-rich loop at the inner mouth of the channel might form a gasket around cargo molecules undergoing export.

August 3, 2017
A flagellum-specific chaperone

Florian's paper on the role of FliO as a flagellum-specific chaperone has been published in PLoS Biology! Congrats!

Florian found that FliO coordinates assembly of the core type-III secretion system (T3SS) of the bacterial flagellum. Upon initiation of flagellum assembly, the flagellar T3SS-specific chaperone FliO facilitates formation of an oligomeric complex of FliP. FliO dissociates from the FliP core complex and is replaced by FliQ and FliR. The FliP/FliQ/FliR complex forms the nucleus for assembly of FlhB and FlhA, followed by MS-ring polymerization and formation of the completed, protein export-competent flagellar T3SS.

June 14, 2017
Microbiota-specific susceptibility towards Salmonella infection

The microbiota composition determines the susceptibility towards Salmonella-induced gastroenteritis. A collaborative work together with Till Strowig at the Helmholtz Centre for Infection Research. Published in Cell Host & Microbe!

Individuals might differ dramatically in their susceptibility to Salmonella infections. To establish a productive infection, Salmonella has to adapt to the intestinal environment and to compete with the host’s microbiota. We hypothesized that individual differences in immune responses and microbiota composition contribute to varying resistance to Salmonella infection and that Salmonella is using metabolic adaptation to overcome these restrictions. In close collaboration with Till Strowig at the Helmholtz Centre for Infection Research, we showed that isogenic mouse lines with distinct microbiota composition differ in susceptibility towards Salmonella infection. The intestinal microbiota ameliorates acute Salmonella-induced diarrhea by priming the immune system and preventing tissue invasion. This is achieved by enhancing antibacterial IFNγ production by innate cells and CD4+ T cells during infection.

March 6, 2017
Mechanism of flagellum growth

Thibaud's paper on the mechanism of filament growth has been published in eLife! Congrats Thibaud!

Thibaud found that bacterial flagella grow through an injection-diffusion mechanism. This flagellum growth model is based on simple biophysical parameters where the filament growth rate is driven by both hindered diffusion and proton motive force-dependent secretion of subunits. The building blocks are pumped into the channel of the flagellum by the type-III secretion system and then travel through diffusion to the tip of the filament. Accordingly, the flagellum grows slower the longer it gets. This molecular mechanism explains also why the growth of bacterial flagella will eventually stop even without any other control mechanisms in place.