Biology subjects

Henke, P.

Publications and source records attributed to Henke, P..

4 recordsLinked to original sources

bioRxiv2026

Interspecies transfer of giant virulence-factor-like proteins in a bacterial symbiosis

The transfer of virulence factors into eukaryotic cells is a hallmark of bacterial pathogenesis. We report the expression, interspecies transfer, subcellular localization, and potential functions of three unusually large virulence factor-like proteins that underlie a bipartite mutualistic bacterial symbiosis. These proteins are synthesized by green sulfur bacterial epibionts surrounding a central motile chemoheterotroph in the multicellular phototrophic consortium Chlorochromatium aggregatum. While symbiosis-proteins remain intracellular during axenic epibiont growth, they are transferred to the partner bacterium in the association. An RTX-like protein secreted towards the central bacterium is capable of degrading its alginate capsule, thereby promoting direct cell-to-cell contact. Two gigantic hemagglutinin-like proteins are predicted to fold when binding extracellular Ca2+ to form Type 6-like auto injection needles, explaining their observed transfer into the central bacterium. These functionalities extend far beyond the known pathogenic interactions of bacteria with eukaryotes and provide new perspectives on the evolution of bacterial virulence factors.

microbiology↗

bioRxiv2023

Isolation and quantification of bacterial membrane vesicles for quantitative metabolic studies using mammalian cell cultures

Bacterial membrane vesicles (BMVs) are produced by most bacteria and participate in various cellular processes, such as intercellular communication, nutrient exchange, and pathogenesis. Notably, these vesicles can contain virulence factors, including toxic proteins, DNA, and RNA. Such factors can contribute to the harmful effects of bacterial pathogens on host cells and tissues. Although the general effects of BMVs on host cellular physiology are well known, the underlying molecular mechanisms are less understood. In this study, we introduce a vesicle quantification method, leveraging the membrane dye FM4-64. We utilize a linear regression model to analyze the fluorescence emitted by stained vesicle membranes to ensure consistent and reproducible vesicle-host interaction studies using cultured cells. This method is particularly valuable for identifying host cellular processes impacted by vesicles and their specific cargo. Moreover, it outcompetes clearly unreliable protein concentration-based methods. We (1) show a linear correlation between the quantity of vesicles and the fluorescence signal emitted from the FM4-64 dye, (2) introduce the "vesicle load" as a new semi-quantitative unit, facilitating more reproducible vesicle-cell culture interaction experiments (3) show that a stable vesicle load yields consistent host responses when studying vesicles from Pseudomonas aeruginosa mutants (4) demonstrate that typical vesicle isolation contaminants, such as flagella, do not significantly skew the metabolic response of lung epithelial cells to P. aeruginosa vesicles, and (5) identify inositol-1-monophosphatase (SuhB) as a pivotal regulator in the vesicle-mediated pathogenesis of P. aeruginosa.

cell biology↗

bioRxiv2020

Functional diversity of isoprenoidal lipids in Methylobacterium extorquens PA1

Hopanoids and carotenoids are two of the major isoprenoid-derived lipid classes in prokaryotes that have been proposed to have similar membrane ordering properties as sterols. Methylobacterium extorquens contains hopanoids and carotenoids in their outer membrane, making them an ideal system to investigate whether isoprenoid lipids play a complementary role in outer membrane ordering and cellular fitness. By genetically knocking out hpnE, and crtB we disrupted the production of squalene, and phytoene in Methylobacterium extorquens PA1, which are the presumed precursors for hopanoids and carotenoids, respectively. Deletion of hpnE unexpectedly revealed that carotenoid biosynthesis utilizes squalene as a precursor resulting in a pigmentation with a C30 backbone, rather than the previously predicted C40 phytoene-derived pathway. We demonstrate that hopanoids but not carotenoids are essential for growth at high temperature. However, disruption of either carotenoid or hopanoid synthesis leads to opposing effects on outer membrane lipid packing. These observations show that hopanoids and carotenoids may serve complementary biophysical roles in the outer membrane. Phylogenetic analysis suggests that M. extorquens may have acquired the C30 pathway through lateral gene transfer with Planctomycetes. This suggests that the C30 carotenoid pathway may have provided an evolutionary advantage to M. extorquens. ImportanceAll cells have a membrane that delineates the boundary between life and its environment. To function properly, membranes must maintain a delicate balance of physical and chemical properties. Lipids play a crucial role in tuning membrane properties. In eukaryotic organisms from yeast to mammals, sterols are essential for assembling a cell surface membrane that can support life. However, bacteria generally do not make sterols, so how do they solve this problem? Hopanoids and carotenoids are two major bacterial lipids, that are proposed as sterol surrogates. In this study we explore the bacterium M. extorquens for studying the role of hopanoids and carotenoids in surface membrane properties and cellular growth. Our findings suggest that hopanoids and carotenoids may serve complementary roles balancing outer membrane properties, and provide a foundation for elucidating the principles of surface membrane adaptation.

microbiology↗

bioRxiv2019

Dinoroseobacter shibae outer membrane vesicles are enriched for the chromosome dimer resolution site dif

Outer membrane vesicles (OMVs) of Gram-negative bacteria have key roles in pathogenesis. However, little is known about their biogenesis and cargo in marine bacteria. In Dinoroseobacter shibae, a marine member of the Rhodobacteraceae, OMVs were produced throughout exponential growth, and DNA could be detected by fluorescence microscopy inside appr. 65% of vesicles. Single cell analysis using time-lapse microscopy showed that individual cells secreted multiple OMVs, preferentially at the septum during cell division. OMVs were enriched for saturated fatty acids, thus their secretion likely increases the fluidity of the membrane of the releasing cell locally. DNA was isolated from the vesicle lumen and sequenced; it was up to 40fold enriched for the region around the terminus of replication (ter). Within this region, the peak of coverage of vesicle DNA was located at dif, a conserved 28 bp palindromic sequence required for binding of the site specific tyrosine recombinases XerCD which are activated by the divisome protein FtsK immediately prior to septum formation. Some of the most abundant proteins of the vesicle proteome were predicted to be required for direct interaction with peptidoglycan during cell division. Single cell analysis, electron microscopy, proteome and DNA cargo show that constitutive OMV secretion in D. shibae occurs mainly prior to septum formation. The footprint of the FtsK/XerCD molecular machinery which resolves chromosome dimers suggests a novel highly conserved route for incorporation of DNA into OMVs. Clearing the division site from small DNA fragments might be an important function of this type of vesicles.

microbiology↗