Biology subjects

Gerlic, M.

Publications and source records attributed to Gerlic, M..

10 recordsLinked to original sources

bioRxiv2026

Choice of host model and manipulated transcription regulator dictates T6SS effector-mediated toxicity

Type VI secretion system (T6SS), a toxic effector-delivery apparatus primarily studied for its antibacterial properties, has recently emerged as a widespread anti-eukaryotic determinant, particularly in members of the genus Vibrio. Although various anti-eukaryotic effectors have been described, it remains unknown whether effectors target distinct hosts in a similar manner. Moreover, it is unclear whether the entire effector arsenal encoded within a single genome is co-regulated under different conditions and by different regulators. Here, we employ the anti-eukaryotic T6SS3 of Vibrio proteolyticus to address these knowledge gaps. By monitoring toxicity during infection of oyster hemocytes and comparing the results with previous infections of murine macrophages, we find that the effector Tie1 plays a role only in intoxicating murine cells, whereas Tie2 intoxicates both cell models. Furthermore, we show that artificial induction of T6SS3 via deletion of the negative regulator hns1 fails to induce the orphan effector Tie3, whereas overexpression of a T6SS3-specific activator, Ats3, induces all three known effectors. Ats3 overexpression further reveals that Tie3, not only Tie2, participates in hemocyte intoxication. Collectively, these findings indicate that analyses of T6SS-mediated infections must factor in the possibility of partial effector repertoire activation even when the main gene cluster is fully induced, and that the contribution of anti-eukaryotic T6SS effectors to intoxication is host-dependent.

microbiology↗

bioRxiv2026

Aeromonas adhesins facilitate kin and non-kin attachment to enable T6SS-mediated antagonism in liquid

Bacterial ability to deploy the type VI secretion system (T6SS) against rivals requires prolonged cell-cell interactions. Such interactions are facilitated on solid surfaces but are assumed to be absent in liquid, leading to the conventional dismissal of T6SS-mediated competition in liquid environments. Here, we find that Aeromonas jandaei employs its T6SS to eliminate diverse bacterial competitors in liquid media. Using a workflow that monitors interbacterial competition via prey luminescence, we demonstrate that auto-aggregation and co-aggregation, facilitated by distinct adhesins, enable kin and non-kin recognition and intoxication in a T6SS-dependent manner. Furthermore, we show that another marine bacterium, Vibrio coralliilyticus, employs T6SS to intoxicate rivals in liquid media. Collectively, our results indicate that T6SS-mediated competition in liquid is more common in marine bacteria than previously anticipated, and can be facilitated by diverse molecular mechanisms that govern cell aggregation.

microbiology↗

bioRxiv2025

Assessing toxicity and competitive fitness of vibrios isolated from coastal waters in Israel

Several species of aquatic bacteria belonging to the genus Vibrio are emerging pathogens of humans and marine animals. Vibrio-associated infections have been shown to correlate with the increase in the oceans surface water temperatures. A slow yet steady increase in vibrios isolated from clinical settings in Israel over the past decade led us to investigate the pathogenic potential of vibrios in Israels coastal waters. We isolated and sequenced the genomes of 23 vibrios from the Mediterranean and the Red Sea. Analysis of these genomes revealed the presence of diverse toxin secretion systems and toxins, as well as mobile genetic elements known to facilitate the dissemination of fitness-enhancing determinants. Moreover, we showed that at least 10 of these isolates can induce cell death in bone marrow-derived macrophages and that at least 12 isolates can intoxicate a rival Vibrio strain in interbacterial competition. Lastly, we determined the susceptibility profiles of these isolates to common antibiotics used to treat Vibrio infections and found widespread resistance to azithromycin. Taken together, our results reveal pathogenic potential within the Vibrio population of Israels coastal waters and underline the need for continued environmental monitoring of emerging pathogens. ImportanceThe oceans surface water temperatures have increased in past decades due to climate change. This increase correlates with the spread of vibrios, a genus of aquatic bacteria, many of which are pathogens of humans and marine animals. Since Vibrio-associated illnesses are rising in Israel, we set out to investigate the Vibrio population in Israels coastal environments and monitor their pathogenic potential. We found diverse repertoires of predicted toxins, the ability to kill mammalian immune cells, and traits that enhance bacterial fitness, such as antibacterial toxicity and resistance to antibiotics commonly used to treat Vibrio infections. These findings indicate that pathogenic traits are circulating within the environmental Vibrio population in Israels coastal waters and suggest that continued monitoring is essential to identify emerging pathogenic strains.

microbiology↗

bioRxiv2024

A T6SS in the coral pathogen Vibrio coralliilyticus secretes an arsenal of anti-eukaryotic effectors and contributes to virulence

Vibrio coralliilyticus (Vcor) is a pathogen of coral and shellfish, leading to devastating economic and ecological consequences worldwide. Although rising ocean temperatures correlate with increased Vcor pathogenicity, the specific molecular mechanisms and determinants contributing to virulence remain poorly understood. Here, we systematically analyzed the type VI secretion system (T6SS), a contact-dependent toxin delivery apparatus, in Vcor. We identified two omnipresent T6SSs that are activated at temperatures in which Vcor becomes virulent; T6SS1 is an antibacterial system mediating interbacterial competition, whereas T6SS2 mediates anti-eukaryotic toxicity and contributes to mortality during infection of an aquatic model organism, Artemia salina. Using comparative proteomics, we identified the T6SS1 and T6SS2 toxin arsenals of three Vcor strains with distinct disease etiologies. Remarkably, T6SS2 secretes at least nine novel anti-eukaryotic toxins comprising core and accessory repertoires. We propose that T6SSs differently contribute to Vcors virulence: T6SS2 plays a direct role by targeting the host, while T6SS1 plays an indirect role by eliminating competitors. Author SummaryCoral reefs are diverse ecosystems providing habitats for various fish, invertebrates, and microorganisms. Climate change, leading to rising ocean water temperatures, correlates with coral bleaching and mass mortality events. An implicated causal agent of coral disease outbreaks is the marine bacterium Vibrio coralliilyticus. Here, we found that two toxin injection systems present in all Vibrio coralliilyticus strains are regulated by temperature; we revealed the toxins that they secrete and their function in competition against rival bacteria and in the intoxication of an animal host. Our findings implicate these systems as previously unappreciated contributors to Vibrio coralliilyticus virulence, illuminating possible targets to treat or prevent coral infection.

microbiology↗

bioRxiv2023

Engineering mice for female-biased progeny without impacting genetic integrity and litter size

The ability to influence the sex ratio of mammalian offspring has applications in agriculture and animal welfare. Here we describe a genetic system in mice that produces predominantly female progeny, reaching approximately ninety percent, without affecting litter size or introducing genetic modifications into the offspring. The system consists of a doxycycline-regulated cassette inserted into the Y chromosome, encoding dCas9 and RNA guides intended to impair the function of Y bearing sperm. While the cassette was originally intended to repress a spermatid maturation gene, our analyses did not detect meaningful changes in gene expression or sperm function that would account for the phenotype. Nevertheless, male mice carrying the cassette consistently produced female biased litters, and the effect was reversed by doxycycline, confirming that it depends on cassette expression. Similar female bias was observed following in vitro fertilization using sperm from transgenic males. These findings demonstrate a reproducible, genetically confined, and conditionally regulated system for sex ratio bias in mice. The strategy may prove adaptable across species, even without a complete mechanistic understanding. Graphical AbstractA single line genetic system producing female biased, genetically unmodified progeny in mice by conditionally impairing Y bearing sperm. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=130 SRC="FIGDIR/small/568055v4_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@5f23b3org.highwire.dtl.DTLVardef@20ed2corg.highwire.dtl.DTLVardef@1b3007dorg.highwire.dtl.DTLVardef@e54cf6_HPS_FORMAT_FIGEXP M_FIG C_FIG One sentence summaryY chromosome manipulation yields mostly female mice, preserving litter size without genetic alteration in females. Significance statementThis study describes a genetic approach for producing predominantly female offspring in mice without altering litter size or transmitting genetic modifications to the progeny. A cassette inserted into the Y chromosome of males induces a strong female bias in offspring through a mechanism that remains unclear. The female offspring do not inherit the transgene and are genetically unmodified. This system provides a simple and conditional platform for sex ratio control in mammals, with potential applications in animal breeding and welfare.

bioengineering↗

bioRxiv2023

A Vibrio T6SS-mediated lethality in a marine animal model

Bacteria belonging to the genus Vibrio include many known and emerging pathogens. Horizontal gene transfer of pathogenicity islands is a major contributor to the emergence of new pathogenic Vibrio strains. In the current report, we use the brine shrimp Artemia salina as a model to show that the marine bacterium Vibrio proteolyticus uses a mobile type VI secretion system, T6SS3, to intoxicate eukaryotic hosts. Two T6SS3 effectors, which were previously shown to induce inflammasome-mediated pyroptotic cell death in mammalian phagocytic cells, contribute to this toxicity. Furthermore, we find a novel, widespread T6SS3 effector that also contributes to the lethality mediated by this system against Artemia salina. Therefore, our results reveal a T6SS that is shared among diverse vibrios and mediates host lethality, indicating that it can lead to the emergence of new pathogenic strains.

microbiology↗

bioRxiv2022

Post-phagocytosis activation of NLRP3 inflammasome by two novel T6SS effectors

The type VI secretion system (T6SS) is used by bacteria to deliver toxic effectors directly into target cells. Most T6SSs mediate antibacterial activities, whereas the potential anti-eukaryotic role of T6SS remains understudied. Here, we found a Vibrio T6SS that delivers two novel effectors into mammalian host immune cells. We showed that these effectors induce a pyroptotic cell death in a phagocytosis-dependent manner; we identified the NLRP3 inflammasome as being the underlying mechanism leading to the T6SS-induced pyroptosis. Moreover, we identified a compensatory T6SS-induced pathway that is activated upon inhibition of the canonical pyroptosis pathway. Genetic analyses revealed possible horizontal spread of this T6SS and its anti-eukaryotic effectors into emerging pathogens in the marine environment. Our findings reveal novel T6SS effectors that activate the host inflammasome and possibly contribute to virulence and to the emergence of bacterial pathogens.

immunology↗

bioRxiv2020

Proteomic analysis of necroptotic extracellular vesicles

Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.

cell biology↗

bioRxiv2019

Vibrio pore-forming leukocidin activates pyroptotic cell death via the NLRP3 inflammasome

Cell death mechanisms are central to combat infectious microbes and to drive pathological inflammation. One such mechanism, the inflammasome, controls infection through either activation of caspase-1 and the subsequent secretion of the mature pro-inflammatory cytokine, interleukin 1{beta} (IL-1{beta}), or by stopping the dissemination of intracellular pathogens by inducing pyroptotic cell death in infected cells. Hemolysins, which are pore-forming toxins (PFTs), target the host cell plasma membrane by producing pores with different diameters. These pores alter the permeability of the target membrane, often leading to cell death. We previously discovered a functional and potent pore-forming, leukocidin domain-containing hemolysin produced by the Gram-negative marine bacterium Vibrio proteolyticus (V. proteolyticus), termed VPRH. Although leukocidin domains are found in other known PFTs, VPRH constitutes a distinct, understudied class within the leukocidin superfamily. Since PTFs of other pathogens were shown to induce cell death by activating the inflammasome pathway, we hypothesized that VPRH-induced cell death is mediated by direct activation of the inflammasome in mammalian immune host cells. Indeed, we found that VPRH induced a two-step cell death in primary macrophages. The first, a rapid step, was mediated by activating the NLRP3 inflammasome, leading to caspase-1 activation and GSDMD cleavage that resulted in IL-1{beta} secretion and pyroptotic cell death. The second step was independent of the inflammasome; however, its mechanism remains unknown. This study sets the foundation for better understanding the immunological consequences of inflammasome activation by a new leukocidin class of toxins.

immunology↗

bioRxiv2019

A genetic system for generating single-sex progeny in mice

The ability to preselect the sex of livestock is economically beneficial and significantly increases the welfare and proper use of animals. In the poultry industry, for example, almost all males are brutally and unnecessarily killed shortly after hatching. The labor and associated costs of separation of females from males, as well as the massive killing of males, could be reduced by using a system that biases the sex of the progeny. Here, we provide a first proof of concept for such a system by crossing two genetically engineered mouse lines. The maternal line encodes a functional Cas9 protein on an autosomal chromosome, whereas the paternal line encodes guide RNAs on the Y chromosome targeting vital mouse genes. After fertilization, the presence of both the Y-encoded guide RNAs from the paternal sperm and the Cas9 protein from the maternal egg target the vital genes in males. We show that this breeding consequently self-destructs solely the males. Our results pave the way for a biased sex production of livestock, thus saving labor, costs, and eliminating substantial animal suffering.

developmental biology↗