Biology subjects

Rojas, D.

Publications and source records attributed to Rojas, D..

7 recordsLinked to original sources

bioRxiv2026

Linking immunity and nutrition: Vital DNA extracellular trap formation in Dictyostelium discoideum reveals an ancestral strategy for microbial management

Extracellular traps (ETs) were originally described in neutrophils as DNA-based structures that immobilize microbes and contribute to innate immunity. Subsequent studies revealed that ET formation occurs across diverse immune cell types and can proceed through non-lytic mechanisms involving mitochondrial DNA release. Whether ETosis also operates outside classical immune contexts and what its ancestral functions may be remain incompletely understood. Here, we show that vegetative, phagocytic cells of the social amoeba Dictyostelium discoideum, a professional bacterial predator with phagocytic mechanisms conserved with those of mammalian innate immune cells, deploy extracellular DNA traps in response to bacterial cues. ET formation is selectively induced by specific lipopolysaccharide variants, is not triggered by canonical neutrophil NET inducers, and occurs through a vital ETosis mechanism that preserves membrane integrity and feeding capacity. Ultrastructural analyses provide the first visualization of extracellular traps in Amoebozoa, revealing extracellular filamentous networks that physically capture bacteria. Molecular characterization demonstrates that amoeboid ETs are enriched in mitochondrial DNA and harbor a dynamic proteomic repertoire dominated by mitochondrial components, DNA-associated proteins, and multiple antibacterial effectors. Notably, ET composition varies with the bacterial stimulus, indicating that ETs are not static structures but rather responsive extracellular assemblies. Together, these findings establish ET formation as a regulated response in a unicellular phagocyte and suggest that extracellular traps may have originally functioned in microbial management during feeding, prior to their elaboration as immune effectors in multicellular organisms. HighlightsO_LIVegetative Dictyostelium discoideum cells deploy mitochondrial DNA-based extracellular traps in response to bacterial cues. C_LIO_LIET formation occurs through a vital, non-lytic mechanism that preserves membrane integrity and feeding capacity. C_LIO_LIExtracellular traps exhibit stimulus-dependent composition and are enriched in mitochondrial and antimicrobial proteins. C_LIO_LIETosis functions as an ancestral strategy for microbial containment and management beyond canonical immune contexts. C_LI

microbiology↗

bioRxiv2025

Expanding the toolbox for hypervirulent Klebsiella pneumoniae using the social amoeba Dictyostelium discoideum as a virulence biosensor

Hypervirulent Klebsiella pneumoniae (hvKp) represents a critical global threat due to its capacity to cause severe, community-acquired infections in healthy individuals and its convergence with multidrug resistance. However, current methods to evaluate hvKp virulence are limited by their reliance on mammalian models, which are costly, ethically constrained, and unsuitable for high-throughput applications. Here, we introduce a biosensing approach using the social amoeba Dictyostelium discoideum as a living phenotypic sensor for hvKp virulence. Through a suite of quantitative assays integrating predation resistance, social development, and real-time single-cell motility tracking, we show that D. discoideum can discriminate avirulent, virulent, and hypervirulent K. pneumoniae strains. Using the model hvKp strain SGH10 and its capsule-null mutant {Delta}wcaJ, we demonstrate that capsule production is strongly associated with phagocytosis resistance, developmental arrest, and amoebae paralysis, all quantifiable through imaging and motility-based readouts. Multivariate data integration revealed a clear segregation of virulence phenotypes, validating D. discoideum as a sensitive, scalable, and genetically tractable biosensor for functional virulence detection. This work establishes a conceptual and methodological framework for developing cell-based biosensors that report virulence levels through measurable host behavioral responses and outlines a calibration path to vertebrate endpoints. These systems provide a low-cost alternative to mammalian systems, especially for early-stage evaluation, and a potential foundation for automated high-throughput screening and surveillance of virulence in critical priority bacterial pathogens.

microbiology↗

bioRxiv2023

Dictyostelium discoideum-assisted pharmacognosy of plant resources for discovering antivirulence molecules targeting Klebsiella pneumoniae

The rise of antibiotic-resistant bacterial strains poses a significant global health challenge, underscoring the critical need for innovative strategies to address this threat. Natural products and their derivatives have emerged as a promising reservoir for drug discovery. The social amoeba Dictyostelium discoideum is an advantageous model organism in this effort. Using this invertebrate model, we introduce a novel perspective to screen natural plant extracts for molecules with potential antivirulence activity. As a proof of concept, we established a simple high-throughput assay to screen for antivirulence molecules targeting Klebsiella pneumoniae among extracts of Helenium aromaticum. Thus, we aimed to identify compounds attenuating K. pneumoniae virulence without inducing cytotoxic effects on amoeba cells. Notably, the methanolic root extract of H. aromaticum but not other extracts fulfilled these prerequisites. Further analysis via UHPLC-ESI-MS/MS led to the identification of 24 chemical compounds boasting potential antivirulence attributes. This research underscores the potential of employing D. discoideum-assisted pharmacognosy for unearthing novel antivirulence agents against multidrug-resistant pathogens. Table of Content Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/564015v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@17530faorg.highwire.dtl.DTLVardef@e28259org.highwire.dtl.DTLVardef@dbc87eorg.highwire.dtl.DTLVardef@147de2d_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2023

Effects of Polyphosphate metabolism Mutations on Biofilm, Capsule Formation and Virulence Traits in Hypervirulent ST23 Klebsiella pneumoniae SGH10

The emergence of hypervirulent Klebsiella pneumoniae (hvKP) strains poses a significant threat to public health due to their high mortality rates and propensity to cause severe community-acquired infections in otherwise healthy individuals. The ability of hvKP to form biofilms and produce a protective capsule contributes to its enhanced virulence and is a significant challenge to effective antibiotic treatment. Therefore, understanding the molecular mechanisms underlying hvKP virulence and biofilm formation is crucial for developing new therapeutic strategies. Polyphosphate Kinase 1 (PPK1) is an enzyme responsible for inorganic polyphosphate synthesis and plays a vital role in regulating various physiological processes in bacteria. In this study, we investigated the impact of polyP metabolism on the biofilm and capsule formation and virulence traits in hvKP using Dictyostelium discoideum amoeba as a model host. We found that the PPK1 null-mutant was impaired in biofilm and capsule formation and showed attenuated virulence in D. discoideum compared to the wild-type strain. We performed a shotgun proteomic analysis of the PPK1 mutant and wild-type strain to gain further insight into the underlying molecular mechanism. The results revealed that the PPK1 mutant had a differential expression of proteins (DEP) involved in capsule synthesis (Wzi - Ugd), biofilm formation (MrkC-D-H), synthesis of the colibactin genotoxin precursor (ClbB), as well as proteins associated with the synthesis and modification of lipid A (ArnB -LpxC - PagP). These proteomic findings corroborate the phenotypic observations and indicate that the PPK1 mutation is associated with impaired biofilm and capsule formation and attenuated virulence in hypervirulent K. pneumoniae. Overall, our study highlights the importance of polyP synthesis in regulating extracellular biomolecules and virulence in K. pneumoniae and provides insights into potential therapeutic targets for treating K. pneumoniae infections.

microbiology↗

bioRxiv2023

MiR-324-5p regulates the structure of dendritic spines and is essential for hippocampal long-term potentiation

MicroRNAs are an emerging class of synaptic regulators. These small noncoding RNAs post-transcriptionally regulate gene expression, thereby altering neuronal pathways and shaping cell-to-cell communication. Their ability to rapidly alter gene expression and target multiple pathways makes them interesting candidates in the study of synaptic plasticity. Here, we demonstrate that the proconvulsive microRNA miR-324-5p regulates excitatory synapse structure and function in the hippocampus of mice. Both Mir324 knockout (KO) and miR-324-5p antagomir treatment significantly reduce dendritic spine density in the hippocampal CA1 subregion, and Mir324 KO, but not miR-324-5p antagomir treatment, shift dendritic spine morphology, reducing the proportion of thin, "unstable" spines. Western blot and quantitative Real-Time PCR revealed changes in protein and mRNA levels for potassium channels, cytoskeletal components, and synaptic markers, including MAP2 and Kv4.2, which are essential for long-term potentiation (LTP). In line with these findings, slice electrophysiology revealed that LTP is severely impaired in Mir324 KO mice, while baseline excitatory activity remains unchanged. Overall, this study demonstrates that miR-324-5p regulates dendritic spine density, morphology, and plasticity in the hippocampus, potentially via multiple cytoskeletal and synaptic modulators.

neuroscience↗

bioRxiv2023

Chiropterans are a hotspot for horizontal transfer of DNA transposons in Mammalia

Horizontal transfer of transposable elements is an important mechanism contributing to genetic diversity and innovation. Bats (order Chiroptera) have repeatedly been shown to experience horizontal transfer of transposable elements at what appears to be a high rate compared to other mammals. We investigated the occurrence of horizontally transferred DNA transposons involving bats. We found over 200 putative horizontally transferred elements within bats; sixteen transposons were shared across distantly related mammalian clades and two other elements were shared with a fish and two lizard species. Our results indicate that bats are a hotspot for horizontal transfer of DNA transposons. These events broadly coincide with the diversification of several bat clades, supporting the hypothesis that DNA transposon invasions have contributed to genetic diversification of bats.

molecular biology↗

bioRxiv2017

Phylogenetic analyses reveal an increase in the speciation rate of raphid pennate diatoms in the Cretaceous

The raphid pennates (order Bacillariales) are a diverse group of diatoms easily recognized by having a slit in the siliceous cell wall, called the raphe, with functions in cell motility. It has been hypothesized that this morphological innovation contributed to the evolutionary success of this relatively young but species-rich group of diatoms. However, owing to the incompleteness of the fossil record this hypothesis remains untested. Using the 18S ribosomal RNA gene, fossil calibrations, and Bayesian phylogenetic and diversification frameworks, we detect a shift in the speciation rate of marine raphid pennate diatoms in the Cretaceous, not detected in other diatom lineages nor previously recognized in the microfossil record. Our results suggest a positive link between the speciation of raphid pennate diatoms and the benefits derived from evolving motility skills, which could account for their outstanding present-day global diversity. The coincidence between the advent of the raphe and the increase in the speciation rate of raphid pennates supports the idea that simple morphological novelties can have important consequences on the evolutionary history of eukaryotic microorganisms.

ecology↗