Biology subjects

Thormann, K. M.

Publications and source records attributed to Thormann, K. M..

7 recordsLinked to original sources

bioRxiv2024

Role of a single MCP in evolutionary adaptation of Shewanella putrefaciens for swimming in planktonic and structured environments

Bacteria can adapt to their environments by changing phenotypic traits by mutations. However, improving one trait often results in deterioration of another one, a trade-off which limits the degree of adaption. The gammaproteobacterium Shewanella putrefaciens CN-32 has an elaborate motility machinery comprising two distinct flagellar systems and an extensive chemotaxis array with 36 methyl-accepting chemotaxis sensor proteins (MCPs). In this study we performed experimental selection on S. putrefaciens for increased spreading through a porous environment. We readily obtained a mutant that showed a pronounced increase in covered distance. This phenotype was almost completely caused by a deletion of 24 bp from the chromosome, which leads to a moderately enhanced production of a single MCP. Accordingly, chemotaxis assays under planktonic conditions and cell tracking in soft agar showed that the mutation improved navigation through nutritional gradients. The study demonstrates how differences in the abundance of a single MCP can lead to an efficient upgrade of directed flagella-mediated motility in specific environments at a low expense of cellular resources. ImportanceExperimental evolution experiments have been used to determine the trade-offs occurring in specific environments. Several studies that have used the spreading behavior of bacteria in structured environments identified regulatory mutants that increase the swimming speed of the cells. While this results in a higher chemotaxis drift, the growth fitness decreases as the higher swimming speed requires substantial cellular resources. Here we show that rapid chemotaxis adaptation can also be achieved through modification of the chemotaxis signal input at a low metabolic cost for the cell.

microbiology↗

bioRxiv2024

Identifying the components of the Shewanella phage LambdaSo lysis system

Phage-induced lysis of Gram-negative bacterial hosts usually requires a set of phage lysis proteins, a holin, an endopeptidase and a spanin system, to disrupt each of the three cell envelope layers. Genome annotations and previous studies identified a gene region in the Shewanella oneidensis prophage LambdaSo, which comprises potential holin- and endolysin-encoding genes but lacks an obvious spanin system. By a combination of candidate approaches, mutant screening, characterization and microscopy we found that LambdaSo uses a pinholin/signal-anchor-release (SAR) endolysin system to induce proton-leakage and degradation of the cell wall. Between the corresponding genes we found that two extensively nested open reading frames encode a two-component spanin module Rz/Rz1. Unexpectedly, we identified another factor strictly required for LambdaSo-induced cell lysis, the phage protein Lcc6. Lcc6 is a transmembrane protein of 65 amino acid residues with hitherto unknown function, which acts at the level of holin in the cytoplasmic membrane to allow endolysin release. Thus, LambdaSo-mediated cell lysis requires at least four protein factors (pinholin, SAR-endolysin, spanin, Lcc6). The findings further extend the known repertoire of phage proteins involved in host lysis and phage egress. SignificanceFor the release of the assembled virions, phages have to breach the cell envelope. For Gram-negatives, this requires the disruption of three layers, the outer and inner membrane and the cell wall. In most cases, the lysis systems of phages infecting Gram-negatives comprises holins to disrupt or depolarize the membrane, thereby releasing or activating endolysins, which then degrade the cell wall. This, in turn, allows the spanins to become active and fuse outer and inner membrane, completing cell envelope disruption and allowing phage egress. Here we show that the presence of these three components may not be sufficient to allow cell lysis, implicating that also in known phages further factors may be required.

microbiology↗

bioRxiv2023

A rolling circle-replicating plasmid as an Inovirus phage satellite

Bacterial viruses (phages) are potent agents of lateral gene transfer and thus are important drivers of evolution. A group of mobile genetic elements (MGEs), referred to as phage satellites, exploit phages to disseminate their own genetic material. Here we isolated a novel member of the genus Inovirus, Shewanella phage Dolos, along with an autonomous rolling circle-replicating plasmid, pDolos. Dolos causes a chronic infection in its host Shewanella oneidensis by phage production with only minor effects on the host cell proliferation. When present, plasmid pDolos hijacks Dolos functions to be predominantly packaged into phage virions and released into the environment. pDolos can disseminate further genetic material encoding, e.g., resistances, fluorophores, and metabolically active proteins, to host cells sensitive to Dolos infection. Given the rather simple requirements of a plasmid for takeover of an inovirus, the wide distribution of phages of this group and the broad spectrum of rolling circle-replicating plasmids, we speculate that similar phage-satellite systems are common among bacteria.

microbiology↗

bioRxiv2023

A conserved cell-pole determinant organizes proper polar flagellum formation

The coordination of cell cycle progression and flagellar synthesis is a complex process in motile bacteria. In {gamma}-proteobacteria, the localization of the flagellum to the cell pole is mediated by the SRP-type GTPase FlhF. However, the mechanism of action of FlhF, and its relationship with the cell pole landmark protein HubP remain unclear. In this study, we discovered a novel protein called FipA that is required for normal FlhF activity and function in polar flagellar synthesis. We demonstrated that membrane-localized FipA interacts with FlhF and is required for normal flagellar synthesis in Vibrio parahaemolyticus, Pseudomonas putida, and Shewanella putrefaciens, and it does so independently of the polar localization mediated by HubP. FipA exhibits a dynamic localization pattern and is present at the designated pole before flagellar synthesis begins, suggesting its role in licensing flagellar formation. This discovery provides insight into a new pathway for regulating flagellum synthesis and coordinating cellular organization in bacteria that rely on polar flagellation and FlhF-dependent localization.

microbiology↗

bioRxiv2022

Constitutive production of flagellar proteins is required for proper flagellation in Shewanella putrefaciens

Flagella are multiprotein complexes whose assembly and positioning requires complex spatiotemporal control. Flagellar assembly is thought to be controlled by several transcriptional tiers, which is mediated through various master regulators. Here, we revisited the regulation of flagellar genes in polarly flagellated gammaproteobacteria by the regulators FlrA, RpoN ({sigma}54) and FliA ({sigma}28) in Shewanella putrefaciens CN-32 at the transcript and protein level. As expected, strict control at both levels occurred for for highly abundant flagellar proteins, including the building blocks for the outer rings, rod, hook and filament. In contrast, a number of regulatory and structural proteins were always present also in the absence of the main regulators. Initiation of flagella assembly and motor activation likely relies on the abundance control of only few structural key components required for formation of the MS- and C-ring and the flagellar type III secrection system. We identified {sigma}70-dependent promoters driving constitutive expression of some flagellar genes including the regulators of flagellar number and positioning, FlhF and FlhG. Reduction of the constitutive expression levels resulted in emergence of hyperflagellation. Thus, basal expression and presence of flagellar proteins is required for proper flagellation, which adds a deeper layer to the regulation of flagellar synthesis and assembly. SignificanceThe tier-based transcriptional regulation underlying bacterial flagella synthesis is - with certain variations - well-established in various species. Here we show that initiation and proceeding of flagellar synthesis can be simply based on the control of some key components and highly abundant building blocks. We further identified a 'tier zero, a set of constitutively produced flagellar regulators and building blocks, which is required, for example, to maintain the flagellar counter. We expect this not only to apply to our model species Shewanella, but also to other flagella regulation systems in bacteria.

microbiology↗

bioRxiv2021

A GGDEF domain serves as a spatial on-switch for a phosphodiesterase by direct interaction with a polar landmark protein

In bacteria, the monopolar localization of enzymes and protein complexes can result in a bi-modal distribution of enzyme activity between the dividing cells and heterogeneity of cellular behaviors. In Shewanella putrefaciens, the multidomain hybrid diguanylate cyclase/phosphodiesterase PdeB, which degrades the secondary messenger c-di-GMP, is located at the flagellated cell pole. Here we show how PdeB polar recruitment is mediated by direct interaction between the inactive diguanylate cyclase (GGDEF) domain of PdeB and the C-terminal FimV domain of the polar landmark protein HubP. We demonstrate that this interaction is crucial for full function of PdeB as a phosphodiesterase. Thus, the GGDEF domain serves as a spatially controlled on-switch that effectively restricts PdeBs activity to the flagellated cell pole. We further show that PdeB regulates abundance and activity of at least two crucial surface-interaction factors, the BpfA surface adhesion protein and the MSHA type IV pilus. The heterogeneity in c-di-GMP concentrations that is generated by differences in abundance and temporal polar appearance of PdeB as well as by bi-modal distribution after cell fission orchestrates the population behavior with respect to cell-surface interaction and environmental spreading. SignificancePhenotypic heterogeneity benefits the proliferation of microbial populations in changing environments. Such heterogeneity can be created by recruitment of enzymatic activity to specific cellular compartments, e.g., the cell pole. Here we show how a GGDEF domain of a multidomain phosphodiesterase has adopted the function as a spatial on-switch that is specifically activated upon direct interaction with a polar landmark protein.

microbiology↗

bioRxiv2018

γ-proteobacteria eject their polar flagella under nutrient depletion, retaining flagellar motor relic structures

Bacteria switch only intermittently to motile planktonic lifestyles under favourable conditions. Under chronic nutrient deprivation, however, bacteria orchestrate a switch to stationary phase, conserving energy by altering metabolism and stopping motility. About two-thirds of bacteria use flagella to swim, but how bacteria deactivate this large-molecular machine remains poorly studied. Here we describe the previously unreported ejection of polar sodium-driven motors by {gamma}-proteobacteria. We show that these bacteria eject their flagella at the base of the flagellar hook when nutrients are depleted, leaving a relic of a former flagellar motor in the outer membrane. Subtomogram averages of the full motor and relic reveal that this is an active process, as a plug protein appears in the relic, likely to prevent leakage across their outer membrane. We show that this is a widespread phenomenon demonstrated by the appearance of relic structures in varied {gamma}-proteobacteria including Plesiomonas shigelloides, Vibrio cholerae, Vibrio fischeri, Shewanella putrefaciens and Pseudomonas aeruginosa.

microbiology↗