Biology subjects

Stutzmann, S.

Publications and source records attributed to Stutzmann, S..

8 recordsLinked to original sources

bioRxiv2025

Structure and Activation Mechanism of a Lamassu Phage Defence System

Lamassu is a diverse family of defence systems that protect bacteria, including pandemic strains of Vibrio cholerae, against phage infection. They target essential cellular processes, aborting infection and preventing phage propagation by terminating the infected host. The mechanisms by which Lamassu efectors are activated when needed and otherwise suppressed are unknown. Here, we present structures of a Lamassu defence system from Salmonella enterica. We show that an oligomerization domain of the nuclease efector, LmuA, is sequestered by two tightly-folded SMC-like LmuB protomers and LmuC. Upon activation, liberated LmuA proteins assemble into a cyclic homo-tetramer, in which two of four nuclease domains are brought into proximity to create an active site capable of cleaving DNA. We propose tetramer formation is likely a one-way switch that establishes a threshold to limit potential spontaneous activation and cell death. Our findings reveal a mechanism of cellular defence, involving liberation and oligomerization of immune efectors, and shed light on how Lamassu systems balance potent immune responses with self-preservation.

microbiology↗

bioRxiv2024

Diverse phage defence systems define West African South American pandemic Vibrio cholerae

Our understanding of the factors underlying the evolutionary success of different lineages of pandemic Vibrio cholerae remains incomplete. Interestingly, two unique genetic signatures define the West African South American (WASA) lineage of V. cholerae responsible for the 1991-2001 Latin American cholera epidemic. Here we show these signatures encode diverse anti-phage defence systems. Firstly, the WASA-1 prophage encodes a 2-gene abortive-infection system WonAB that renders the lineage resistant to the major predatory vibriophage ICP1, which alongside other phages, is thought to restrict cholera epidemics and has potential for use in prophylaxis. Secondly, a unique set of genes on the Vibrio seventh pandemic island II encodes an unusual modification-dependent restriction system targeting phages with modified genomes, and a new member of the Shedu defence family that defends against vibriophage X29. Taken together, we propose that these anti-phage defence systems have likely contributed to the success of a major epidemic lineage of the ongoing seventh cholera pandemic.

microbiology↗

bioRxiv2024

Molecular mechanism of plasmid elimination by the DdmDE defense system

Seventh pandemic Vibrio cholerae strains contain two hallmark pathogenicity islands that encode the DNA defense modules DdmABC and DdmDE. Here we use cryo-EM to reveal the mechanistic basis for plasmid defense by DdmDE. A cryo-EM structure of the DdmD helicase-nuclease reveals that it adopts an auto-inhibited dimeric architecture. The prokaryotic Argonaute protein DdmE uses a DNA guide to target plasmid DNA. A structure of the DdmDE complex, validated by in vivo mutational studies, shows that DNA binding by DdmE triggers disassembly of the DdmD dimer and loading of monomeric DdmD onto the non-target DNA strand. Finally, in vitro studies reveal that DdmD translocates in the 5 to 3 direction, while partially degrading the plasmid DNA. These findings provide critical insights into the architecture and mechanism of DdmDE systems in plasmid elimination.

microbiology↗

bioRxiv2022

A single nucleotide polymorphism determines constitutive versus inducible type VI secretion in Vibrio cholerae

Vibrio cholerae is a well-studied human pathogen that is also a common inhabitant of marine habitats. In both environments, the bacterium is subject to interbacterial competition. A molecular nanomachine that is often involved in such competitive behavior is the type VI secretion system (T6SS). Interestingly and in contrast to non-pandemic or environmental isolates, the T6SS of the O1 El Tor clade of V. cholerae, which is responsible for the ongoing 7th pandemic, is largely silent under standard conditions. Instead, these strains induce their full T6SS capacity only under specific conditions such as growth on chitinous surfaces (signaled through TfoX and QstR) or when the cells encounter low intracellular c-di-GMP levels (TfoY-driven). In this study, we identified a single nucleotide polymorphism (SNP) within an intergenic region of the major T6SS gene cluster of V. cholerae that determines the T6SS status of the cell. We show that SNP conversion is sufficient to induce T6SS production in numerous pandemic strains, while the converse approach renders non-pandemic/environmental V. cholerae strains T6SS-silent. We further demonstrate that SNP-dependent T6SS production occurs independently of the known T6SS regulators TfoX, QstR, and TfoY. Finally, we identify a putative promoter region adjacent to the identified SNP that is required for all forms of T6SS regulation.

microbiology↗

bioRxiv2019

Neighbor predation linked to natural competence fosters the transfer of large genomic regions in Vibrio cholerae

Natural competence for transformation is a primary mode of horizontal gene transfer (HGT). Competent bacteria are able to absorb free DNA from their surroundings and exchange this DNA against pieces of their own genome when sufficiently homologous. And while it is known that transformation contributes to evolution and pathogen emergence in bacteria, there are still questions regarding the general prevalence of non-degraded DNA with sufficient coding capacity. In this context, we previously showed that the naturally competent bacterium Vibrio cholerae uses its type VI secretion system (T6SS) to actively acquire DNA from non-kin neighbors under chitin-colonizing conditions. We therefore sought to further explore the role of the T6SS in acquiring DNA, the condition of the DNA released through T6SS-mediated killing versus passive cell lysis, and the extent of the transfers that occur due to these conditions. To do this, we herein measured the frequency and the extent of genetic exchanges in bacterial co-cultures on competence-inducing chitin under various DNA-acquisition conditions. We show that competent V. cholerae strains acquire DNA fragments with an average and maximum length exceeding 50 kbp and 150 kbp, respectively, and that the T6SS is of prime importance for such HGT events. Collectively, our data support the notion that the environmental lifestyle of V. cholerae fosters HGT and that the coding capacity of the exchanged genetic material is sufficient to significantly accelerate bacterial evolution.\n\nSignificance StatementDNA shuffled from one organism to another in an inheritable manner is a common feature of prokaryotes. It is a significant mechanism by which bacteria acquire new phenotypes, for example by first absorbing foreign DNA and then recombining it into their genome. In this study, we show the remarkable extent of the exchanged genetic material, frequently exceeding 150 genes in a seemingly single transfer event, in Vibrio cholerae. We also show that to best preserve its length and quality, bacteria mainly acquire this DNA by killing adjacent, healthy neighbors then immediately absorbing the released DNA before it can be degraded. These new insights into this prey-killing DNA acquisition process shed light on how bacterial species evolve in the wild.

microbiology↗

bioRxiv2018

DNA-uptake pilus of Vibrio cholerae capable of kin-discriminated auto-aggregation

Natural competence for transformation is a widely used and key mode of horizontal gene transfer that can foster rapid bacterial evolution. Competent bacteria take-up DNA from their environment using Type IV pili, a widespread and multi-purpose class of cell surface polymers. However, how pili facilitate DNA-uptake has remained unclear. Here, using direct labelling, we show that in the Gram-negative pathogen Vibrio cholerae DNA-uptake pili are highly dynamic and that they retract prior to DNA-uptake. Unexpectedly, these pili can self-interact to mediate auto-aggregation of cells into macroscopic structures. This phenotype is conserved in disease causing pandemic strains. However, extensive strain-to-strain variability in the major pilin subunit PilA, present in environmental isolates, controls the ability of pili to interact without affecting transformation. We go on to show that interactions between pili are highly specific, enabling cells producing pili composed of different PilA subunits to discriminate between one another. On chitin surfaces, a natural habitat of V. cholerae, pili connect cells within dense networks, suggesting a model whereby DNA-uptake pili function to promote inter-bacterial interactions during surface colonisation. Moreover, our results provide evidence that type IV pili could provide a simple and potentially widespread mechanism for bacterial kin recognition.

microbiology↗

bioRxiv2018

QstR-dependent regulation of natural competence and type VI secretion in Vibrio cholerae

During growth on chitinous surfaces in its natural aquatic environment Vibrio cholerae develops natural competence for transformation and kills neighboring non-immune bacteria using a type VI secretion system (T6SS). Activation of these two phenotypes requires the chitin-induced regulator TfoX, but also integrates signals from quorum sensing via the intermediate regulator QstR, which belongs to the LuxR-type family of regulators. Here, we define the QstR regulon using RNA sequencing. Moreover, by mapping QstR binding sites using chromatin immunoprecipitation coupled with deep sequencing we demonstrate that QstR is likely a dual transcription factor that binds upstream of the up- and down-regulated genes. Like other LuxR-type family transcriptional regulators we show that QstR function is dependent on dimerization. However, in contrast to the well-studied LuxR-type biofilm regulator VpsT of V. cholerae, which requires the second messenger c-di-GMP, we show that QstR dimerization and function is c-di-GMP independent. Surprisingly, although ComEA, which is a periplasmic DNA-binding protein essential for transformation, is produced in a QstR-dependent manner, QstR-binding was not detected upstream of comEA suggesting the existence of a further regulatory pathway. Overall these results provide detailed insights into the function of a key regulator of natural competence and type VI secretion in V. cholerae.

microbiology↗

bioRxiv2017

Molecular insights into Vibrio cholerae’s intra-amoebal host-pathogen interactions

Vibrio cholerae, which causes the diarrheal disease cholera, is a species of bacteria commonly found in aquatic habitats. Within such environments, the bacterium must defend itself against predatory protozoan grazers. Amoebae are prominent grazers, with Acanthamoeba castellanii being one of the best-studied aquatic amoebae. We previously showed that V. cholerae resists digestion by A. castellanii and establishes a replication niche within the hosts osmoregulatory organelle. In this study, we deciphered the molecular mechanisms involved in the maintenance of V. choleraes intra-amoebal replication niche and its ultimate escape from the succumbed host. We demonstrated that minor virulence features important for disease in mammals, such as extracellular enzymes and flagellum-based motility, play a key role role in the replication and transmission of V. cholerae in its aqueous environment. This work, therefore, describes new mechanisms that provide the pathogen with a fitness advantage in its primary habitat, which may have contributed to the emergence of these minor virulence factors in the species V. cholerae.

microbiology↗