Biology subjects

Boon, M.

Publications and source records attributed to Boon, M..

10 recordsLinked to original sources

bioRxiv2026

Genomic Potential vs. Pathogenic Reality: Functional Virulence Variation in Pseudomonas aeruginosa

Despite the rapid expansion of bacterial genome sequencing, predicting pathogen virulence from genomic information remains a major challenge. To study this association, we integrate comparative genomics, functional phenotyping, host-pathogen interaction assays, and infection outcomes across a diverse collection of 28 Pseudomonas aeruginosa isolates, representing clinically relevant diversity in serotype, type IV pilus group, and flagellar type. While virulence-associated systems were broadly retained across the strain collection, isolates displayed substantial variation in phenotypic behavior and pathogenicity. Genotypic classifications based on pilus, flagellar, siderophore, or secretion system characteristics were poor predictors of virulence-associated phenotypes. Pathogenicity in a Galleria mellonella model varied markedly among strains and was most strongly associated with surface-associated motility, whereas other virulence traits and quorum-sensing genotypes showed limited predictive value. In a three-dimensional lung epithelial cell model, inflammatory responses were negatively correlated with activity of the type III secretion system effector ExoS, suggesting a role in modulation of host immune signaling. Together, these findings demonstrate that virulence emerges from functional deployment of conserved virulence repertoires, rather than their mere presence. By linking genomic diversity to phenotypic expression, pathogenicity, and host responses within a single strain collection, this study highlights fundamental limitations of gene-content-based virulence prediction and underscores the importance of integrating genomic and functional data to better understand P. aeruginosa virulence.

microbiology↗

bioRxiv2026

High-resolution transcriptome analysis reveals a multilayered and dynamic transcriptional architecture in the archaeal virus SSV1

Fuselloviridae infecting thermoacidophilic Sulfolobales are among the best-studied archaeal viruses, with the Saccharolobus Spindle Shaped Virus 1 (SSV1)-Saccharolobus solfataricus interaction serving as a model system. After infection, SSV1 establishes a chronic infection with new virions continuously budding from the infected cell. UV irradiation triggers virus replication, increased virion production and a tightly orchestrated temporal transcription cycle. However, gene regulatory mechanisms post-UV-induction remain elusive. In this study, we investigated the SSV1 transcriptomic landscape post-UV-induction using high-resolution, long-read ONT-cappable-RNA-sequencing. Through end-to-end sequencing of primary transcripts, we generated a comprehensive transcript map of SSV1, identifying 21 transcription start sites (TSSs), including 12 newly described sites, and 19 termination sites (TTSs), of which 9 are novel. This enabled refinement of transcription units and operon structures across the genome. Five new antisense RNAs were discovered, including one encoded antisense to the open reading frame of a putative viral toxin gene a291, which we hypothesize to be a cis-encoded regulatory RNA. In addition, spontaneous recombination between two direct repeats in the SSV1 structural genes vp1 and vp3 was confirmed. To our knowledge, this is the first implementation of cappable-seq for an archaeal virus, and the first base-resolution analysis of the SSV1 transcriptome. It reveals a previously unappreciated level of transcriptional complexity, illustrating how high-resolution transcriptomics can deepen understanding of viral transcriptome architecture and regulatory networks. ImportanceArchaeal viruses remain among the least understood viruses in the virosphere, despite their ecological and evolutionary significance. Saccharolobus Spindle Shaped Virus 1 (SSV1) is a key model for the study of virus-host interactions in archaea. Yet, its gene regulatory mechanisms have remained poorly resolved. Here, we provide the first base-pair resolution transcriptome of an archaeal virus, revealing a complex and multi-layered transcriptional architecture. We uncover alternative transcription start and termination sites, as well as antisense transcription, pointing to regulatory mechanisms beyond a simple temporal gene expression program. These findings suggest that archaeal viruses use sophisticated transcriptional and post-transcriptional mechanisms similar to those in bacterial and eukaryotic viruses. By establishing a high-resolution map of the SSV1 transcriptome, this work advances our understanding of archaeal virus biology.

microbiology↗

bioRxiv2026

Polyamine dysregulation converges with RASopathies on RAS/MAPK and sensory processing phenotypes in Drosophila

RASopathies are developmental conditions associated with cognitive and sensory processing impairments. They are caused by pathogenic variants in genes that result in overactivation of the RAS/MAPK signaling pathway. Genes linked to this pathway have been reported to be enriched among Drosophila models with habituation deficits, a behavioral phenotype reflecting sensory filtering. To identify hidden RASopathies - monogenic disorders that converge on RAS/MAPK overactivation without being classically linked to the pathway - we generated 89 and screened 41 viable habituation-deficient Drosophila RNAi models for RAS/MAPK overactivation, measured as an increased phosphorylated ERK to ERK ratio. This screen identified Sms, the ortholog of human spermine synthase (SMS), implicated in Snyder-Robinson syndrome. RAS/MAPK overactivation along with hyperreactivity and habituation impairments are confirmed in a full loss-of-function mutant. A RNAi screen targeting polyamine pathway genes identified Sat (human SAT1/2, SATL1) to reproduce these phenotypes. Knockdown of Sms or Sat in GABAergic neurons impaired habituation, implicating polyamine metabolism in inhibitory circuit function. These findings reveal previously unrecognized convergence between polyamine dysregulation and RASopathies, suggesting shared therapeutic opportunities through modulation of either pathway. SUMMARY STATEMENTUsing Drosophila, we uncovered polyamine metabolism genes, Sms and Sat, as modulators of RAS/MAPK and sensory processing, revealing a shared mechanism between polyaminopathies and RASopathies that may inform unified therapies.

genetics↗

bioRxiv2023

Refining the transcriptional landscapes for distinct clades of virulent phages infecting Pseudomonas aeruginosa

The introduction of high-throughput sequencing has resulted in a surge of available bacteriophage genomes, unveiling their tremendous genomic diversity. However, our current understanding of the complex transcriptional mechanisms that dictate their gene expression during infection is limited to a handful of model phages. Here, we applied ONT-cappable-seq to reveal the transcriptional architecture of six different clades of virulent phages infecting Pseudomonas aeruginosa. This long-read microbial transcriptomics approach is tailored to globally map transcription start and termination sites, transcription units and putative RNA-based regulators on dense phage genomes. Specifically, the full-length transcriptomes of LUZ19, LUZ24, 14-1, YuA, PAK_P3 and giant phage phiKZ during early, middle and late infection were collectively charted. Beyond pinpointing traditional promoter and terminator elements and transcription units, these transcriptional profiles provide insights in transcriptional attenuation and splicing events and allow straightforward validation of Group I intron activity. In addition, ONT-cappable-seq data can guide genome-wide discovery of novel regulatory element candidates, including non-coding RNAs and riboswitches. This work substantially expands the number of annotated phage-encoded transcriptional elements identified to date, shedding light on the intricate and diverse gene expression regulation mechanisms in Pseudomonas phages, which can ultimately be sourced as tools for biotechnological applications in phage and bacterial engineering.

microbiology↗

bioRxiv2023

Validation of the sleep EEG headband ZMax

Polysomnography (PSG) is the gold standard for recording sleep. However, the standard PSG systems are bulky, expensive, and often confined to lab environments. These systems are also time-consuming in electrode placement and sleep scoring. Such limitations render standard PSG systems less suitable for large-scale or longitudinal studies of sleep. Recent advances in electronics and artificial intelligence enabled wearable PSG systems. Here, we present a study aimed at validating the performance of ZMax, a widely-used wearable PSG that includes frontal electroencephalography (EEG) and actigraphy but no submental electromyography (EMG). We analyzed 135 nights with simultaneous ZMax and standard PSG recordings amounting to over 900 hours from four different datasets, and evaluated the performance of the headbands proprietary automatic sleep scoring (ZLab) alongside our open-source algorithm (DreamentoScorer) in comparison with human sleep scoring. ZLab and DreamentoScorer compared to human scorers with moderate and substantial agreement and Cohens kappa scores of 59.61% and 72.18%, respectively. We further analyzed the competence of these algorithms in determining sleep assessment metrics, as well as shedding more lights on the bandpower computation, and morphological analysis of sleep microstructural features between ZMax and standard PSG. Relative bandpower computed by ZMax implied an error of 5.5% (delta), 4.5% (theta), 1.6% (alpha), 0.5% (sigma), 0.8% (beta), and 0.2% (gamma), compared to standard PSG. In addition, the microstructural features detected in ZMax did not represent exactly the same characteristics as in standard PSG. Besides similarities and discrepancies between ZMax and standard PSG, we measured and discussed the technology acceptance rate, feasibility of data collection with ZMax, and highlighted essential factors for utilizing ZMax as a reliable tool for both monitoring and modulating sleep.

neuroscience↗

bioRxiv2022

Tail-tape-fused virion and non-virion RNA polymerases of a thermophilic virus with an extremely long tail

Thermus thermophilus bacteriophage P23-45 encodes a giant 5,002-residue tail tape measure protein (TMP)1 that defines the length of its extraordinarily long 800 nm tail2,3. We found that the N-terminal portion of P23-45 TMP is an unusual RNA polymerase (RNAP) homologous to cellular and viral two-barrel RNAPs. The TMP-fused virion RNAP transcribes pre-early phage genes, including a gene that encodes another, non-virion RNAP, that transcribes early and some middle phage genes. We determined the crystal structures of both P23-45 RNAPs. The non-virion RNAP has a crab claw-like architecture similar to previously reported two-barrel RNAPs. The virion RNAP adopts a unique flat structure without a clamp, which likely reflects the requirement for its extrusion through the narrow channel in the phage tail for delivery into the cell. Structure and sequence comparisons of the P23-45 RNAPs with other phage and cellular RNAPs suggest that, despite the extensive functional differences, the two P23-45 RNAPs originate from an ancient gene duplication in an ancestral phage. Our findings demonstrate remarkable adaptability of two-barrel RNAPs that can be attained within a single virus species.

molecular biology↗

bioRxiv2022

Cold exposure induces dynamic changes in circulating triacylglycerol species, which is dependent on intracellular lipolysis: a randomized cross-over trial

BackgroundThe application of cold exposure has emerged as an approach to enhance whole-body lipid catabolism. The global effect of cold exposure on the lipidome in humans has been reported with mixed results depending on intensity and duration of cold. MethodsThis secondary study was based on data from a previous randomized cross-over trial (ClinicalTrials.gov ID: NCT03012113). We performed sequential lipidomic profiling in serum during 120 min cold exposure of human volunteers. Next, the intracellular lipolysis was blocked in mice using a small-molecule inhibitor of adipose triglyceride lipase (ATGL; Atglistatin), and were exposed to cold for a similar duration. The quantitative lipidomic profiling was assessed in-depth using the Lipidyzer platform. FindingsCold exposure gradually increased circulating free fatty acids reaching a maximum at 60 min, and transiently decreased total triacylglycerols (TAGs) only at 30 min. A broad range of TAG species was initially decreased, in particular unsaturated and polyunsaturated TAG species with [≤]5 double bonds, while after 120 min a significant increase was observed for polyunsaturated TAG species with [≥]6 double bonds. The mechanistic study in mice revealed that the cold-induced increase in polyunsaturated TAGs was largely prevented by blocking adipose triglyceride lipase. InterpretationWe interpret these findings as that cold exposure feeds thermogenic tissues with TAG-derived fatty acids for combustion, resulting in a decrease of circulating TAG species, followed by increased hepatic production of polyunsaturated TAG species induced by liberation of free fatty acids stemming from adipose tissue. Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSCold exposure has emerged as a novel non-pharmacological strategy to enhance whole-body lipid catabolism, to improve lipid homeostasis and ultimately cardiometabolic health. In mice, cold exposure accelerates the clearance of triacylglycerol (TAG)-rich lipoproteins from the circulation, reducing circulating TAGs. In humans however, the effect of cold exposure on whole-body TAG metabolism remained thus far controversial, as mixed results are reported depending on intensity and duration of cold. Added value of this studyHere, we performed sequential lipidomic profiling in serum during 120 min cold exposure of human volunteers. We show that cold exposure gradually increases circulating free fatty acids and transiently decreases total TAGs after 30 min, accompanied with a dynamic change in circulating TAGs as dependent on their saturation status and length. Specifically, cold exposure decreases a broad range of more saturated and shorter TAG species, while after 90-120 min polyunsaturated TAG species with [≥]6 double bonds start to increase. Subsequently, we performed a mechanistic study in mice, in which we show that the cold-induced increase in polyunsaturated TAGs is largely prevented when blocking intracellular lipolysis. Implications of all the available evidenceOur findings describe a mechanism by which cold exposure provides thermogenic tissues with TAG-derived fatty acids for combustion. At the same time, cold exposure increases lipolysis in white fat to drive hepatic TAG production to further feed thermogenic tissues. For the first time, these results show that the TAG lowering effect of cold exposure as observed in mice can be recapitulated in humans, which warrants further studies on the beneficial effects of cold exposure on accelerating lipid metabolism to improve cardiometabolic health.

physiology↗

bioRxiv2022

Transcriptomics-driven characterisation of novel T7-like temperate Pseudomonas phage LUZ100

The Autographiviridae is a diverse yet distinct family of bacterial viruses marked by a strictly lytic lifestyle and a generally conserved genome organization. We here characterise Pseudomonas aeruginosa phage LUZ100, a distant relative of type phage T7. LUZ100 is a podovirus with a limited host range and identified LPS as the likely phage receptor. Interestingly, infection dynamics of LUZ100 indicated moderate adsorption rates and low virulence, hinting towards temperate behavior. This hypothesis was supported by genomic analysis, which showed that LUZ100 shares the conventional T7-like genome organization, yet encodes key genes associated with a temperate lifestyle. To unravel the peculiar characteristics of LUZ100, ONT-cappable-seq transcriptomics analysis was performed. This data generated a birds-eye view of the LUZ100 transcriptome and enabled the discovery of key regulatory elements, antisense RNA, and transcriptional unit structures. The transcriptional map of LUZ100 also allowed us to identify new RNAP-promoter pairs that can form the basis for biotechnological parts and tools for new synthetic transcription regulation circuitry. The ONT-cappable-seq data revealed that the LUZ100 integrase and a MarR-like regulator (proposed to be involved in the lytic/lysogeny decision), are actively co-transcribed in an operon. In addition, the presence of a phage-specific promoter transcribing the phage-encoded RNA polymerase, raises questions on the regulation of this polymerase, and suggests it is interwoven with the MarR-based regulation. This transcriptomics-driven characterisation of LUZ100 supports the increasing evidence that T7-like phages should not straightforwardly be marked as having a strictly lytic lifecycle. ImportanceBacteriophage T7, considered the model phage of the Autographiviridae family, is marked by a strictly lytic lifecycle and conserved genome organisation. Recently, novel phages of this clade are emerging and showing characteristics associated to a lysogenic lifecycle. Screening for temperate behaviour is of outmost importance in fields like phage therapy, where strictly lytic phages are generally required for therapeutic applications. In this study, weve used an omics-driven approach to characterise the T7-like Pseudomonas aeruginosa phage LUZ100. These results led to the identification of actively transcribed lysogeny-associated genes in the phage genome, pointing out that temperate T7-like phages are emerging more frequent than initially thought. In short, the combination of genomics and transcriptomics allowed us to obtain a better understanding of the biology of non-model Autographiviridae phages, which can be used to optimize the implementation of phages and their regulatory elements in phage therapy and biotechnological applications, respectively.

microbiology↗

bioRxiv2022

Development of ONT-cappable-seq to unravel the transcriptional landscape of Pseudomonas phages

RNA sequencing has become the method of choice to study the transcriptional landscape of phage-infected bacteria. However, short-read RNA sequencing approaches generally fail to capture the primary 5 and 3 boundaries of transcripts, confounding the discovery of key transcription initiation and termination events as well as operon architectures. Yet, the elucidation of these elements is crucial for the understanding of the strategy of transcription regulation during the infection process, which is currently lacking beyond a handful of model phages. To this end, we developed ONT-cappable-seq, a specialized long-read RNA sequencing technique that allows end-to-end sequencing of primary prokaryotic transcripts using the Nanopore sequencing platform. We applied ONT-cappable-seq to study transcription of Pseudomonas aeruginosa phage LUZ7, obtaining a comprehensive genome-wide map of viral transcription start sites, terminators, and complex operon structures that fine-regulate gene expression. Our work provides new insights in the RNA biology of a non-model phage, unveiling distinct promoter architectures, putative small non-coding viral RNAs, and the prominent regulatory role of terminators during infection. The robust workflow presented here offers a framework to obtain a global, yet fine-grained view of phage transcription and paves the way for standardized, in depth transcription studies for microbial viruses or bacteria in general.

genomics↗

bioRxiv2018

Differential Antiviral Activities of RSV Inhibitors in Human Airway Epithelium

We report the use of reconstituted 3D-human airway epithelium cells of bronchial origin (HuAEC) in an air-liquid interface to study respiratory syncytial virus (RSV) infection and to assess the efficacy of RSV inhibitors in (pre-)clinical development. RSV-A replicates efficiently in HuAEC and viral RNA is shed for weeks after infection. RSV infection reduces the ciliary beat frequency of the ciliated cells as of 4 days post infection, with complete ciliary dyskinesia observed by day 10. Treatment with RSV fusion inhibitors resulted in an antiviral effect only when added at the time of infection. In contrast, the use of replication inhibitors (both nucleoside and non-nucleosides) elicited a marked antiviral effect even when start of treatment was delayed until one or even three days after infection. Levels of the inflammation marker RANTES (mRNA) increased [~]200-fold in infected-untreated cultures (at three weeks post infection), but levels were comparable to those of uninfected cultures in the presence of PC-876, a RSV-replication inhibitor, demonstrating that an efficient antiviral treatment inhibits virus induced inflammation in this model. Overall, HuAEC offer a robust and physiologically relevant model to study RSV replication and to assess the efficacy of antiviral compounds.

microbiology↗