Biology subjects

Klavins, K.

Publications and source records attributed to Klavins, K..

10 recordsLinked to original sources

bioRxiv2026

NeuroEnergetics-on-Chip: a novel 3-compartment microfluidic platform to study metabolic interactions between brain parenchyma and cerebral vasculature

Neurological disorders are a major cause of death and disability worldwide. The brains energy metabolism is essential to its proper function, yet the mechanisms driving neuroenergetic dysfunction remain poorly understood. A key challenge is the limited availability of human-relevant models that can reproduce the complexity of brain physiology. An Organ-on-Chip (OoC) system was developed to mimic the neurovascular unit metabolic coupling by incorporating human isogenic iPSC-derived endothelial-like cells, pericyte-like cells, astrocytes, and a cerebral organoid, representing the main cellular components of the NVU. The novel, customized microfluidic platform enables research on neurovascular coupling by interconnecting a blood-brain barrier-on-a-chip model with a 3D brain parenchymal compartment to mimic physiological conditions. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/733618v1_ufig1.gif" ALT="Figure 1"> View larger version (75K): org.highwire.dtl.DTLVardef@9fc9f3org.highwire.dtl.DTLVardef@838a7corg.highwire.dtl.DTLVardef@1f8df76org.highwire.dtl.DTLVardef@16ef40_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

pH-dependent antibacterial activity of N-acetylcysteine against Staphylococcus aureus and metabolic alterations induced at neutral pH

N-acetylcysteine (NAC) is a mucolytic and antioxidant increasingly investigated as an antibacterial agent and antibiotic adjuvant, particularly against biofilm-associated infections. Despite being a weak organic acid, solution pH is rarely reported in the literature, and reported minimum inhibitory concentrations (MIC) for S. aureus are highly inconsistent across studies, varying by more than 50-fold. We systematically assessed the pH-dependence of NAC antibacterial activity and investigated how pH-neutral NAC perturbs bacterial metabolism using LC-MS based targeted metabolomics. Without pH-adjustment, NAC at 200 mM (pH 2.8) acted bactericidal against the S. aureus strains USA300 and Mu12, while pH-adjusted NAC solutions had no effect on bacterial growth, even at concentrations approaching its solubility limit. Based on LC-MS measurement, pH adjustment did not measurably degrade NAC but significantly increased dimerization ratios, suggesting that altered ionization state rather than degradation underlies the loss of antibacterial activity at neutral pH. Despite the absence of growth inhibition, pH-neutral NAC induced concentration-dependent metabolic alterations under both planktonic and biofilm conditions. Arginine, cysteine, glycolysis, and TCA cycle were the most affected pathways, with intracellular accumulation of arginine and cystine, together with increased lactic acid production. Our findings demonstrate that the antibacterial activity of NAC against S. aureus is driven by acidification, helping reconcile contradictory MIC reports in the literature. Additionally, pH-neutral NAC alters bacterial metabolism without impairing growth, highlighting its potential to modulate bacterial physiology independently of direct antibacterial activity.

microbiology↗

bioRxiv2026

Gallium induces cytotoxicity through disruption of DNA synthesis rather than ferroptosis

BackgroundGallium (Ga) is a promising anti-tumor agent; however, its precise molecular targets in osteosarcoma remain debated. While current paradigms largely attribute its toxicity to reactive oxygen species (ROS) and ferroptosis, understanding its true mechanism is essential for overcoming therapeutic resistance. This highlights the need for interdisciplinary approaches, such as metabolomics, to unveil novel vulnerabilities in cancer metabolism. MethodsWe employed an interdisciplinary strategy utilizing high-resolution liquid chromatography-mass spectrometry (LC-MS) metabolomics and 13C2-glutamine stable isotope tracing in osteosarcoma cells to elucidate the cytotoxic mechanisms of gallium nitrate. Scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM-EDS) was utilized for elemental mapping, and in silico modeling was applied to evaluated metal binding dynamics. Furthermore, synergistic effects were tested by combining gallium with the DNA-damaging agent cisplatin. ResultsOur metabolic profiling revealed a profound bifurcation characterized by the systemic depletion of glycolysis and pentose phosphate pathway intermediates, coupled with a novel ribonucleotide accumulation bottleneck. The observed distinct signature strongly implicated ribonucleotide reductase (RNR) as the primary enzymatic target. In silico modeling and SEM-EDS visually and thermodynamically confirmedthat gallium acts as a structural decoy for iron within the RNR active site. The co-localization induces functional iron starvation rather than canonical ferroptosis. Furthermore, isotope tracing confirmed that elevated ROS is a consequence of overall metabolic failure, not the primary driver of cell death. Crucially, gallium functioned as a metabolic DNA repair inhibitor, synergizing potently with cisplatin to prevent the repair of platinum-induced DNA lesions. ConclusionsGallium selectively sensitizes highly proliferative sarcoma cells by disrupting RNR-mediated DNA precursor synthesis, while sparing normal osteoblasts. Leveraging metabolomics to uncover this state of functional iron starvation provides a rational, interdisciplinary framework for developing gallium-based combination therapies designed to break platinum resistance in clinical oncology.

cancer biology↗

bioRxiv2025

Modulation of Neutrophil Function by Foetal Bovine Serum and Type I Collagen

Immune regulation plays a crucial role during the regeneration process, and it determines the fate of inflammation after tissue injury or infection. Neutrophils serve as the primary immune cells recruited to the site of inflammation, initiating and directing the subsequent inflammatory cascade following implantation. As neutrophils are highly sensitive to environmental cues, commonly used culture supplements and molecules may themselves influence neutrophil function and consequently affect the interpretation of biomaterial-induced immune responses. This study examined how foetal bovine serum (FBS), a standard in vitro culture supplement, affects human peripheral blood neutrophils when supplied either in the culture medium or as a surface coating on 3D-printed polycaprolactone (PCL) scaffolds. The response to type I collagen coating was also assessed as a biologically relevant comparator. Neutrophil activity was evaluated by assessing metabolic activity and metabolomic profiles, reactive oxygen species (ROS) production, and inflammation-related markers via a high-throughput proximity extension assay. Type I collagen coating modified the metabolomic profile of neutrophils and MMP-9 release but had minimal effect on ROS generation. In contrast, the presence of FBS in the culture medium significantly influenced neutrophil behavior, leading to significant changes in metabolic activity, cytotoxicity, and the secretion of inflammation-associated molecules, even at concentrations as low as 1% (v/v). These findings highlight the importance of assessing the use of FBS in neutrophil culture models, particularly those isolated from humans, and emphasize the need to develop alternative platforms for investigating neutrophil-biomaterial interactions in a more physiologically relevant manner.

immunology↗

bioRxiv2025

Purine nucleobases enhance CD8+ T cell effector function

During antiviral immune responses, activated immune cells remodel metabolic pathways towards uptake and utilization of biosynthetic and bioenergetic metabolites. Concurrently, viral infections alter metabolic environments, impacting metabolite availability for the establishment of an effective immune response. Here, we integrated in vivo metabolomics data from murine and human viral infections with in vitro metabolite screens, identifying purine nucleobases as novel immunometabolites that enhance CD8+ T cell effector function. We found that CD8+ T cells can switch from resource-intensive purine de novo synthesis to purine salvage pathway, to produce nucleotides from purine nucleobases. This strategy of metabolic adaptation allows diversion of biosynthetic and bioenergetic resources towards enhancing effector molecule production. Our findings unveil an adaptation strategy by CD8+ T cells to manage metabolic challenges in dynamic organismal environments and suggest pharmacological targets in purine metabolism as potential targets for immunotherapy. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=144 SRC="FIGDIR/small/669899v1_ufig1.gif" ALT="Figure 1"> View larger version (65K): org.highwire.dtl.DTLVardef@17b4b15org.highwire.dtl.DTLVardef@17e57a3org.highwire.dtl.DTLVardef@1abc36aorg.highwire.dtl.DTLVardef@1aa704a_HPS_FORMAT_FIGEXP M_FIG C_FIG Instead of producing nucleotides via purine de novo synthesis, CD8+ T cells can import and utilize purine nucleobases via the purine salvage pathway to divert bioenergetic and biosynthetic resources towards effector function. By shifting from purine de novo synthesis to the purine salvage pathway, cells save significant resources: 5 moles of the key bioenergetic metabolite ATP, and biosynthetic metabolites including 2 moles of glutamine, 1 mole each of serine or glycine, and 1 mole of aspartate.

immunology↗

bioRxiv2025

Functional profiling reveals a non-enzymatic role of NUDT5 in repressing purine de novo synthesis

Folate metabolism is intricately linked to purine de novo synthesis through the incorporation of folate-derived one-carbon units into the purine scaffold. Here, we investigate the chemical and genetic dependencies caused by mutations in the folate enzyme MTHFD1 and discover a key role for Nudix hydrolase 5 (NUDT5) in regulating purine de novo synthesis. Through genetic knockout and development of a selective chemical NUDT5 degrader, we uncover an unprecedented scaffolding role rather than NUDT5 enzymatic activity is responsible for this phenotype. We find that NUDT5 interacts with the rate-limiting enzyme of purine de novo synthesis, PPAT, to repress the pathway in response to elevated purine levels. Our findings establish NUDT5 as an important regulator of purine de novo synthesis and elucidate its role in mediating sensitivities to 6-thioguanine in cancer treatment and to adenosine in MTHFD1 deficiency.

molecular biology↗

bioRxiv2024

Metabolic mapping of the human solute carrier superfamily

Solute carrier (SLC) transporters govern most of the chemical exchange across cellular membranes and are integral to metabolic regulation, which in turn is linked to cellular function and identity. Despite their key role, individual functions of the members of the SLC superfamily were not evaluated systematically. We determined the metabolic and transcriptional profiles upon SLC overexpression in knock-out or wild-type isogenic cell backgrounds. Targeted metabolomics provided a fingerprint of 189 intracellular metabolites, while transcriptomics offered insights into cellular programs modulated by SLC expression. Beyond the metabolic profiles of 102 SLCs directly related to their known substrates, we also identified putative substrates or metabolic pathway connections for 71 SLCs without previously annotated bona fide substrates, including SLC45A4 as a new polyamine transporter. By comparing the molecular profiles, we identified functionally related SLC groups, including some with distinct impacts on osmolyte balancing and glycosylation. The assessment of functionally related human genes presented here may serve as a blueprint for other systematic studies of human gene function and supports future investigations into the functional roles of SLCs.

systems biology↗

bioRxiv2022

Gain-of-function genetic screens in human cells identify SLC transporters overcoming environmental nutrient restrictions

Solute carrier (SLC) transporters control fluxes of nutrients and metabolites across membranes and thereby represent a critical interface between the microenvironment and cellular and subcellular metabolism. Because of substantial functional overlap, the interplay and relative contributions of members of this family in response to environmental stresses remain poorly elucidated. In order to infer functional relationships between SLCs and metabolites, we developed a strategy to identify human SLCs able to sustain cell viability and proliferation under growth-limiting concentrations of essential nutrients. One-by-one depletion of 13 amino acids required for cell proliferation enabled gain-of-function genetic screens using a SLC-focused CRISPR/Cas9-based transcriptional activation approach to uncover transporters relieving cells from the growth-limiting metabolic bottleneck. We identified the cationic amino acid transporter SLC7A3 as a gene that, when upregulated, overcame low availability of arginine and lysine by increasing their uptake. SLC7A5 (LAT1), on the other hand, was able to sustain cellular fitness upon deprivation of several neutral amino acids. A genome-wide screen identified SLC7A3 as the single main gene product able to rescue cell survival in the limiting arginine conditions tested, demonstrating the potentially decisive role of transporters in overcoming nutrient limitations. Moreover, we identified metabolic compensation mediated by the glutamate/aspartate transporters SLC1A2 and SLC1A3 under glutamine-limiting conditions. Overall, this gain-of-function approach using human cells led to the definition of functional transporter-nutrient relationships and revealed that upregulation of transport activity may be sufficient to overcome environmental metabolic restrictions.

cell biology↗

bioRxiv2019

Ifnar1 signaling breaks the hepatic urea cycle to regulate adaptive immunity

Infections induce complex host responses linked to antiviral defense, inflammation and tissue damage and repair. These processes are increasingly understood to involve systemic metabolic reprogramming. We hypothesized that the liver as a central metabolic hub may orchestrate many of these changes during infection. Thus, we investigated the systemic interplay between inflammation and metabolism in a mouse model of chronic viral infection and hepatitis. Here we show that virus-induced type I interferon (IFN-I) modulates wide-spread metabolic alterations of the liver in a hepatocyte-intrinsic Ifnar1-dependent way. Specifically, IFN-I repressed the transcription of numerous genes with metabolic function including Otc and Ass1, which encode enzymes of the urea cycle. This led to decreased arginine and increased ornithine concentrations in the circulation, resulting in suppressed virus-specific CD8 T cell responses and ameliorated liver pathology. These findings establish IFN-I-induced modulation of hepatic metabolism and the urea cycle as an endogenous mechanism of immunoregulation.

immunology↗

bioRxiv2018

MTHFD1 is a genetic interactor of BRD4 and links folate metabolism to transcriptional regulation

The histone acetyl-reader BRD4 is an important regulator of chromatin structure and transcription, yet factors modulating its activity have remained elusive. Here we describe two complementary screens for genetic and physical interactors of BRD4, which converge on the folate pathway enzyme MTHFD1. We show that a fraction of MTHFD1 resides in the nucleus, where it is recruited to distinct genomic loci by direct interaction with BRD4. Inhibition of either BRD4 or MTHFD1 results in similar changes in nuclear metabolite composition and gene expression, and pharmacologic inhibitors of the two pathways synergize to impair cancer cell viability in vitro and in vivo. Our finding that MTHFD1 and other metabolic enzymes are chromatin-associated suggests a direct role for nuclear metabolism in the control of gene expression.

cancer biology↗