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RESEARCH

In order to read more about a research project of your choice, simply click on the corresponding picture.

 

Cytokinesis and morphogenesis

Cytokinesis is a prerequisite for cellular life. Therefore, it is not surprising that the process of cell division is tightly controlled and intimately linked to other key cellular processes such as genome replication. Understanding the molecular mechanisms behind cytokinesis revealed fascinating insights in the subcellular organization of eukaryotic and prokaryotic cells.

 

Our lab is devoted to understand how Gram positive, rod-shaped bacteria, such as Bacillus subtilis and Corynebacterium glutamicum, are accomplishing cell division and how this process is regulated in time and space. Currently, we focus on function of the Min system in B. subtilis and division site selection in C. glutamicum. Furthermore, we study cell growth of bacteria using the apically growing C. glutamicum as a model system. C. glutamicum is a close relative of notorious pathogens such as C. diphterieae and Mycobacterium tuberculosis. These bacteria share a complex multilayered cell envelope. We are interested in the machineries that build the cell wall and their the spatiotemporal regulation.

Read more: 

FtsZ induces membrane deformations via torsional stress upon GTP hydrolysis
Ramirez-Diaz DA, Merino-Salomón A, Meyer F, Heymann M, Rivas G, Bramkamp M, Schwille P.
Nat Commun, 12(1):3310. doi: 10.1038/s41467-021-23387-3.

Dynamics of the Bacillus subtilis Min system
Feddersen H, Würthner L, Frey E, Bramkamp M.
mBio, 12(2):e00296-21. doi: 10.1128/mBio.00296-21.

A novel component of the division-site selection system of Bacillus subtilis and a new mode of action for the division inhibitor MinCD
Bramkamp M, Emmins R, Weston L, Donovan C, Daniel RA, Errington J
Mol Microbiol., 2008 Dec;70(6):1556-69.

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Figure 1: Bacillus subtilis cell with FtsZ-GFP (green), nucleoids (blue) and cell membrane (red). Transmission electron micrograph of a dividing Corynebacterium glutamicum cell. Merged image showing a B. subtilis Min mutant strain, expressing FtsZ-GFP (green), nucleoids (red) and phase contrast. Cells show abnormal division and mini-cell formation. (descriptions from left to right)

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Chromosome segregation and organization

The bacterial chromosome exceeds the size of the bacterial cell by several orders of magnitude. Therefore, bacterial genomes are organized in compact nucleoids that contain the highly condensed DNA. The compacted DNA still allows for efficient replication and transcription. A major challenge for the cell is segregation of the replicated DNA into the daughter cells to ensure viable offspring.

 

We study the mechanisms of chromosome segregation and organization mainly in C. glutamicum. These bacteria contain at least two chromosomes stably tethered to the cell poles, making this bacterium diploid. The newly replicated DNA uses the existing nucleoid as a track to segregate with a ParAB system towards midcell. This is a good example that chromosomes are not only storage places for genetic information, but play a structural role in cellular self-organization. 

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Chromosome organization by a conserved condensin–ParB system in the actinobacterium Corynebacterium glutamicum
Böhm K, Giacomelli G, Schmidt A, Imhof A, Koszul R, Marbouty M, Bramkamp M.
Nat Commun, 11(1):1485. doi: 10.1038/s41467–020–15238–4.

Novel Chromosome Organization Pattern in Actinomycetales—Overlapping Replication Cycles Combined with Diploidy 
Böhm K, Meyer F, Rhomberg A, Kalinowski J, Donovan C, Bramkamp M
mBio, 2017 Jun 6. vol. 8 no. 3 e00511-17 doi: 10.1128/mBio.00511-17.

Interlinked Sister Chromosomes Arise in the Absence of Condensin during Fast Replication in B. subtilis
Gruber S, Veening J-W, Bach J, Blettinger M, Bramkamp M, Errington J
Curr. Biol., 2014. doi: 10.1016/j.cub.2013.12.049.

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Figure 2: Corynebacterium glutamicum cells expressing DivIVA-mCherry (red) and the centromere-binding protein ParB-YFP (yellow) reveal polar tethering of ParB-parS nucleoprotein complexes. Deletion of ParB gives rise to anucleate cells. Hi-C contact maps reveal drastic differences between chromosome organization in wild type and ParB deletion strains. Single-molecule localization microscopy reveals parS dependency of ParB cluster formation. (descriptions from left to right)

 

Membrane dynamics

Biological membranes are the essential barrier that enable cellular life. One third of all proteins are membrane proteins and help to organize the transfer of information and material from the cytoplasm to the environment. Amazingly, the phospholipid bilayer is highly organized in space and time. It is well established that bacterial protein complexes and lipids are not uniformly distributed in membranes. Proteins and lipids can interact to perform several cellular processes such as signal transduction and protein secretion.

 

Our lab has been pioneering research on bacterial flotillin proteins. Flotillins fluidize membranes and function in membrane homeostasis. This in turns contributes to cell shape maintenance, correct cell wall synthesis and cytoskeleton dynamics. We use several fluorescent probes and fluorescence microscopy techniques to investigate the molecular mechanisms associated with membrane organization and homeostasis regulation in Bacillus subtilis

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Flotillin mediated membrane fluidity controls peptidoglycan synthesis and MreB movement
Zielińska A, Savietto A, de Sousa Borges A, Martinez D, Berbon M, Roelofsen JR, Hartman AM, de Boer R, van der Klei IJ, Hirsch AKH, Habenstein B, Bramkamp M, Scheffers D-J.
Elife, 14;9:e57179. doi: 10.7554/eLife.57179.

Bacterial dynamin-like protein DynA mediates lipid and content mixing
Guo L, Bramkamp M.
FASEB J., 33(11):11746-11757. doi: 10.1096/fj.201900844RR..

Exploring the Existence of Lipid Rafts in Bacteria
Bramkamp M, Lopez D
Microbiol Mol Biol Rev., 2015 Mar;79(1):81-100. doi: 10.1128/MMBR.00036-14.

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Figure 3: The flotillin homolog FloT forms membrane associated cluster in Bacillus subtilis cells (FloT-GFP foci merged on a phase contrast image). Molecular speed of MreB foci measured by TIRF microscopy reveal an influence of flotillins in dynamics of the MreB-governed cell wall synthesis complexes. Kymographs of TIRF microscopy of MreB foci used to calculate the MreB speeds. (descriptions from left to right)

 

Phage biology

Viruses are highly abundant and affect cellular life in various aspects. Viruses that infect and replicate in bacteria are called bacteriophages, or short phages. The exponential replication rates of viral infections are a serious threat to living cells and, hence, several defense systems have been evolved. Most anti-viral systems interfere with phage genome replication and many well-known components that we use for molecular genetics are derived from anti-viral defense mechanisms. This includes restriction endonucleases and CRISPR-Cas systems. Less is known about phage genome dynamics catalyzed by cytoskeletal structures, host cell lysis and phage dispersal. We study phage encoded cytoskeletal proteins, such as the actin homolog AlpC in C. glutamicum. 

A few years ago we discovered a bacterial dynamin-like protein, termed DynA, in B. subtilis. We could show that DynA plays a role in membrane dynamics such as membrane fusion. In vivo DynA is part of the bacterial innate immune systems, because DynA delays host cell lysis after phage replication and prohibits efficient phage dispersal. Thus, DynA has a protective effect on the population level. 

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A dynamin-like protein involved in bacterial cell membrane surveillance under environmental stress
Sawant P, Eissenberger K, Karier L, Mascher T, Bramkamp M
Environ Microbiol, (2016) 18: 2705–2720. doi:10.1111/1462-2920.13110

A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria
Donovan C, Heyer A, Pfeifer E, Polen T, Wittmann A, Krämer R, Frunzke J, Bramkamp M
Nucleic Acids Res., 2015 Apr 27. pii: gkv374. doi: 10.1093/nar/gkv374

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Figure 4: Negative stain electron micrograph of a SPbeta phage (Siphoviridae) and phi29 (Podoviridae). Plaque assay plate of Bacillus subtilis strains infected with phi29. Deletion of the bacterial dynamin-like protein DynA renders cells more sensitive against infection, while overexpression makes cells resistant against phage infection. Fluorescent micrograph showing the infection of B. subtilis with phi29 phages. Phage capsids are labelled in red and viral DNA is labelled in blue. (descriptions from left to right)

 

Antibiotics

Antibiotics are our most powerful weapon against bacterial infections. No other medical invention has had a similar dramatic effect on human life expectancy. However, spreading of antibiotic resistant bacteria becomes a serious problem and need for new antibiotics or new combinations of antibiotic treatments is of utmost importance.

 

Our group tries to understand the cellular effects antibiotics have on bacterial cells to gain deeper understanding in the mechanism of antibiotic action. Precise knowledge on how antibiotics affect bacteria helps to make predictions on how combinations of antibiotics can effectively eradicate bacterial infections. Currently, we study effects of antituberculosis drugs using C. glutamicum as a model system. 

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RNA-mediated control of cell shape modulates antibiotic resistance in Vibrio cholerae
Herzog R, Peschek N, Singh PK, Sprenger M, Meyer F, Fröhlich KS, Schröger L, Bramkamp M, Drescher K, Papenfort K
Nature Commun, 11(1):6067. doi: 10.1038/s41467-020-19890-8

The Antituberculosis Drug Ethambutol Selectively Blocks Apical Growth in CMN Group Bacteria
Schubert K, Sieger B, Meyer F, Giacomelli G, Böhm K, Rieblinger A, Lindenthal L, Sachs N, Wanner G, Bramkamp M
mBio, 2017 Feb 7. vol. 8 no. 1 e02213-16. doi: 10.1128/mBio.02213-16

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Figure 5: Scanning electron micrograph of Corynebacterium glutamicum cells treated with the anti-tuberculosis drug ethambutol. Ethambutol blocks apical, but not septal, cell wall synthesis. C. glutamicum cells expressing DivIVA-mCherry (red) were bio-orthogonally labelled at their sites of nascent cell wall synthesis (green/blue). Checker-board assays are used to show effects of combinatorial treatment with different antibiotics. (descriptions from left to right)

 

Advanced imaging

Our group employs a diverse set of cutting-edge microscopy techniques. Our technical tool box includes fluorescent recovery after photobleaching (FRAP), time-lapse microscopy, (3D) photo-activated localization microscopy, cluster analysis of single molecules, 3D structured illumination and single-particle tracking (SPT) in live-cells. These techniques enable us to visualize cellular structures with nanometer precision. Protein dynamics can be tracked in life cells. Most of these techniques can be applied to a broad spectrum of proteins and organisms and are not limited to bacteria. In the future, we hope our work can help to expand the understanding of essential molecular processes and mechanics like cell division or morphogenesis 

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MamY is a membrane–bound protein that aligns magnetosomes and the motility axis of helical magnetotactic bacteria
Toro–Nahuelpan M, Giacomelli G, Raschdorf O, Borg S, Plitzko JM, Bramkamp M, Schüler D, Müller FD.
Nat Microbiol., 4(11):1978 – 1989. doi: 10.1038/s41564–019–0512–8

Optimization of sample preparation and green color imaging using the mNeonGreen fluorescent protein in bacterial cells for photoactivated localization microscopy
Stockmar I, Feddersen H, Cramer K, Gruber S, Jung K, Bramkamp M, Shin JY
Sci Rep., 2018 Jul 4; 8(1):10137. doi: 10.1038/s41598-018-28472-0

Light Microscopy: Methods and Protocols 
Sample Preparation and Choice of Fluorophores for Single and Dual Color Photo-Activated Localization Microscopy (PALM) with Bacterial Cells
Bach JN, Giacomelli G, Bramkamp M
Methods in Molecular Biology, (2017) vol. 1563

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Figure 6: Single-molecule imaging (SMLM) of a central component of the cytoskeleton in Magnetospirillum gryphiswaldense outlining the geodetic line of the cell. Comparison of conventional wide field imaging and SMLM exemplified using ParB cluster. Single molecule tracking of a membrane integral cell division protein in Corynebacterium glutamicum. (descriptions from left to right)