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Cutruzzola, F.

Publications and source records attributed to Cutruzzola, F..

5 recordsLinked to original sources

Zinc Starvation Drives Respiratory Remodeling and Metabolic Adaptations Associated with Cystic Fibrosis in Pseudomonas aeruginosa

Host nutritional immunity restricts microbial growth by altering metal availability. In patients with cystic fibrosis (CF), microorganisms inhabiting the thick airway mucus experience severe nutrient limitation, particularly zinc (Zn) restriction, which plays a critical role in limiting lung colonization. However, Pseudomonas aeruginosa, a major contributor to morbidity and mortality in CF, employs multiple strategies to overcome Zn deficiency and persist within the airways. Understanding how P. aeruginosa adapts to Zn deprivation may facilitate the development of antimicrobial approaches targeting Zn homeostasis. In this study, we characterized the physiological and transcriptional adaptations that support P. aeruginosa survival under Zn-limited conditions. Transcriptomic analysis of a znuAzrmB mutant unable to efficiently acquire Zn revealed widespread repression of pathways involved in central carbon metabolism, motility and virulence. Notably, Zn limitation promoted extensive respiratory remodeling, characterized by a shift toward anaerobic metabolism, induction of denitrification pathways, altered terminal oxidase expression, and reduced oxygen consumption. These metabolic changes correlated with decreased ATP production, altered membrane potential, and increased aminoglycoside tolerance. Furthermore, the Zn-starved mutant exhibited reduced production of quorum-sensing molecules, redox imbalance and altered oxidative stress responses. Many of these adaptations resemble those observed in P. aeruginosa isolated from CF sputum, suggesting convergence towards a common host-adapted physiological state. Collectively, these findings identify Zn starvation as a major driver of bacterial physiological remodeling in CF conditions and reveal a previously unrecognized link between Zn limitation, respiratory reprogramming, and the emergence of persistence-associated traits in P. aeruginosa. ImportanceDuring infection, the host restricts Zn availability as part of nutritional immunity, but how this influences the physiology of Pseudomonas aeruginosa remains poorly understood. Here we show that severe Zn limitation triggers a coordinated metabolic program that extends far beyond Zn acquisition, encompassing respiratory remodeling, altered energy metabolism, redox imbalance, and increased tolerance to aminoglycosides. Remarkably, many of these changes resemble transcriptional and physiological traits previously described in bacteria adapted to cystic fibrosis airways, identifying Zn limitation as a key environmental signal that contributes to chronic infection-associated phenotypes. These findings broaden our understanding of how metal availability shapes bacterial physiology and suggest that targeting Zn homeostasis may influence both bacterial persistence and antibiotic susceptibility.

microbiology↗

Merging Multi-OMICs with Proteome Integral Solubility Alteration Unveils Antibiotic Mode of Action

Antimicrobial resistance is responsible for an alarming number of deaths, estimated at 5 million per year. To combat priority pathogens, like Helicobacter pylori, the development of novel therapies is of utmost importance. Understanding the molecular alterations induced by medications is critical for the design of multi-targeting treatments capable of eradicating the infection and mitigating its pathogenicity. However, the application of bulk omics approaches for unraveling drug molecular mechanisms of action is limited by their inability to discriminate between target-specific modifications and off-target effects. This study introduces a multi-omics method to overcome the existing limitation. For the first time, the PISA assay is utilized in bacteria in the PISA-express format to link proteome solubility with different and potentially immediate responses to drug treatment, enabling us the resolution to understand target-specific modifications and off-target effects. This study introduces a comprehensive method for understanding drug mechanisms and optimizing the development of multi-targeting antimicrobial therapies.

microbiology↗

THE RIBOREGULATION MECHANISM OF HUMAN SERINE HYDROXYMETHYLTRANSFERASE IS ROOTED IN AN ALLOSTERIC SWITCH

RNA can directly control protein activity in a process called riboregulation; only a few mechanisms of riboregulation have been described in detail, none of these being characterized on structural grounds. Here we present a comprehensive structural, functional, and phylogenetic analysis of riboregulation of cytosolic serine hydroxymethyltransferase (SHMT1), the enzyme interconverting serine and glycine in one-carbon metabolism. We show that the RNA modulator competes with polyglutamylated folates and acts as an allosteric switch, selectively altering the enzymes reactivity vs. serine. In addition, we identify the tetrameric assembly and a flap structural motif as key structural elements necessary for binding of RNA to eukaryotic SHMT1. The results presented here suggest that riboregulation may have played a role in the evolution of eukaryotic SHMT1 and the compartmentalization of one-carbon metabolism. The findings also provide insights for RNA-based therapeutic strategies targeting this cancer-linked metabolic pathway.

biochemistry↗

SHMT1-RNA interaction dynamically regulates serine and glycine concentration in cells

Human serine hydroxymethyltransferase (SHMT) regulates the serine-glycine one carbon metabolism and plays a role in cancer metabolic reprogramming. Two SHMT isozymes are acting in the cell: SHMT1 encoding the cytoplasmic isozyme, and SHMT2 encoding the mitochondrial one. Here we present a molecular model built on experimental data reporting the interaction between SHMT1 protein and SHMT2 mRNA, recently discovered in lung cancer cells. Using a stochastic dynamic model, we show that RNA moieties dynamically regulate serine and glycine concentration, shaping the system behaviour. For the first time we observe an active functional role of the RNA in the regulation of the serine-glycine metabolism and availability, which unravels a complex layer of regulation that cancer cells exploit to fine tune amino acids availability according to their metabolic needs. The quantitative model, complemented by an experimental validation in the lung adeno-carcinoma cell line H1299, exploits RNA molecules as metabolic switches of the SHMT1 activity. Our results pave the way for the development of RNA-based molecules able to unbalance serine metabolism in cancer cells.

biochemistry↗

Cytosolic localization and in vitro assembly of human de novo thymidylate synthesis complex

De novo thymidylate synthesis is a crucial pathway for normal and cancer cells. Deoxythymidine monophosphate (dTMP) is synthesized by the combined action of three enzymes: serine hydroxymethyltransferase (SHMT), dihydrofolate reductase (DHFR) and thymidylate synthase (TYMS), the latter two targets of widely used chemotherapeutics such as antifolates and 5-fluorouracil. These proteins translocate to the nucleus after SUMOylation and are suggested to assemble in this compartment into the thymidylate synthesis complex (dTMP-SC). We report the intracellular dynamics of the complex in lung cancer cells by in situ proximity ligation assay, showing that it is also detected in the cytoplasm. This result strongly indicates that the role of the dTMP-SC assembly may go beyond dTMP synthesis. We have successfully assembled the dTMP synthesis complex in vitro, employing tetrameric SHMT1 and a bifunctional chimeric enzyme comprising human TYMS and DHFR. We show that the SHMT1 tetrameric state is required for efficient complex assembly, indicating that this aggregation state is evolutionary selected in eukaryotes to optimize protein-protein interactions. Lastly, our results on the activity of the complete thymidylate cycle in vitro, may provide a useful tool to develop drugs targeting the entire complex instead of the individual components.

biochemistry↗