Biology subjects

Henriquez, T.

Publications and source records attributed to Henriquez, T..

3 recordsLinked to original sources

bioRxiv2026

MxtR/ErdR is a central regulator of short-chain fatty acid metabolism in Pseudomonas alloputida

Short-chain fatty acids (SCFAs) such as acetate and propionate represent important carbon and energy sources for environmental pseudomonads, requiring coordinated regulation to ensure efficient assimilation while maintaining metabolic homeostasis. In Pseudomonas alloputida KT2440, the two-component system MxtR/ErdR (CrbS/CrbR) is known to activate acetate assimilation through regulation of acsA-I. However, the full extent of the MxtR/ErdR regulon and its physiological role beyond acetate metabolism have remained unclear. We demonstrate that MxtR/ErdR directly regulates the methylcitrate cycle and coordinates both acetate and propionate metabolism. To define the MxtR/ErdR regulatory network, we combined comparative transcriptomics, physiological analyses, promoter-reporter assays, electrophoretic mobility shift assays (EMSAs) and targeted mutagenesis. Comparative transcriptomic analyses of the mxtR-H806N and {Delta}erdR mutants revealed extensive changes in gene expression, including coordinated downregulation of genes involved in propionate metabolism alongside genes associated with central carbon metabolism, transport, chemotaxis, and signal transduction. Consistent with these transcriptional changes, deletion of either mxtR or erdR abolished growth on propionate. Promoter-reporter assays and EMSAs demonstrated direct binding of ErdR to a conserved imperfect inverted repeat upstream of the prp gene cluster and prpE. Mutational analyses confirmed the functional importance of this binding motif for promoter activation. In addition, MxtR/ErdR contributed to pyruvate utilization through regulation of transport-associated genes, whereas deletion of downstream target genes only caused modest phenotypes, indicating that the physiological effects of MxtR/ErdR arise from coordinated regulation of multiple pathways. Our findings substantially expand the MxtR/ErdR regulon and identify this signaling system as a central regulator of SCFA homeostasis in P. alloputida KT2440.

microbiology↗

bioRxiv2025

Establishment and application of a vesicle extraction method for clinical strains of Pseudomonas aeruginosa

Pseudomonas aeruginosa is a versatile pathogen capable of causing illnesses that range from mild infections to life-threatening conditions. Its virulence is driven by a wide array of factors, among which extracellular vesicles (EVs) have gained recognition as important contributors to its pathogenicity. Despite this, the full scope of their roles remains unclear. A major barrier to EV characterization is the difficulty of vesicle isolation--procedures are often lengthy, yield is low, and specialized equipment is required. In this study, we assessed the effectiveness of a rapid vesicle extraction method from clinical strains of P. aeruginosa. To that end, we first selected and characterized six phenotypically diverse clinical strains of P. aeruginosa (two reference strains and 4 clinical isolates, including one strain from a cystic fibrosis patient) and used them to evaluate the vesicle extraction method. The results obtained through SDS-PAGE analysis, western blot, protein quantification, and TEM indicated the presence of vesicles in all samples; however, it was also possible to observe a large number of contaminants in some of them (mainly LS07 and Z37). Subsequent treatment with enzymes (DNase and/or alginate lyase) allowed for the elimination of the contaminants as observed by electron microscopy. Our results suggest that the method is suited for the vesicle extraction of clinical isolates of P. aeruginosa. The phenotypic complexity of these strains presents challenges that current rapid purification methods are ill-equipped to handle, highlighting the need for improved or alternative approaches.

microbiology↗

bioRxiv2024

Branched Oncolytic Peptides Target HSPGs, Inhibit Metastasis, and Trigger the Release of Molecular Determinants of Immunogenic Cell Death in Pancreatic Cancer.

Immunogenic cell death (ICD) can be exploited to treat non-immunoreactive tumors that do not respond to current standard and innovative therapies. Not all chemotherapeutics trigger ICD, among those that do exert this effect, there are anthracyclines, irinotecan, some platinum derivatives and oncolytic peptides. We studied two new branched oncolytic peptides, BOP7 and BOP9 that proved to elicit the release of damage-associated molecular patterns DAMPS, mediators of ICD, in pancreatic cancer cells. The two BOPs selectively bound and killed tumor cells, particularly PANC-1 and Mia PaCa-2, but not cells of non-tumor origin such as RAW 264.7, CHO-K1 and pgsA-745. The cancer selectivity of the two BOPs may be attributed to their repeated cationic sequences, which enable multivalent binding to heparan sulfate glycosaminoglycans (HSPGs), bearing multiple anionic sulfation patterns on cancer cells. This interaction of BOPs with HSPGs not only fosters an anti-metastatic effect in vitro, as demonstrated by reduced adhesion and migration of PANC-1 cancer cells, but also shows promising tumor-specific cytotoxicity and low hemolytic activity. Remarkably, the cytotoxicity induced by BOPs triggers the release of DAMPs, particularly HMGB1, IFN-{beta} and ATP, by dying cells, persisting longer than the cytotoxicity of conventional chemotherapeutic agents such as irinotecan and daunorubicin. An in vivo assay in nude mice showed an encouraging 20% inhibition of tumor grafting and growth in a pancreatic cancer model by BOP9.

biochemistry↗