Biology subjects

Berkers, C. R.

Publications and source records attributed to Berkers, C. R..

10 recordsLinked to original sources

bioRxiv2026

MASH-in-a-dish reveals fatty acid-driven itaconate crosstalk between Kupffer cells and hepatocytes

Metabolic dysfunction-associated steatohepatitis (MASH) is characterized by lipid-driven sterile inflammation and fibrosis that leads to liver failure and liver cancer. Metabolites are increasingly recognized as signaling molecules and regulate disease progression through cell-cell crosstalk. To overcome the challenges of studying metabolite crosstalk between different cell types in MASH livers, we developed a novel in vitro MASH model, "MASH-in-a-dish", a Transwell co-culture system of mouse hepatocyte-derived organoids (HepOrgs), Kupffer cells (KCs), and hepatic stellate cells (HSCs) in a chemically defined liver co-culture medium. MASH-in-a-dish models key clinical features of clinical MASH, including steatosis, hepatocyte ballooning, inflammation and fibrosis. MASH-in-a-dish allowed for rapid and specific separation of co-cultured HepOrgs, KCs and HSCs, enabling cell type-specific metabolomics and stable-isotope fluxomics analysis. Using MASH-in-a-dish, we identified 19 MASH-associated, cell interaction-responsive metabolites and demonstrated that fatty acids, via the carnitine shuttle, drive KC itaconate production, which aggravated oxidative stress, steatosis, and cell damage in lipid-stressed HepOrgs. Inhibiting itaconate production by KC-specific Acod1 knockout in the MASH-in-a-dish system mitigated hepatocyte damage. Our results show that MASH-in-a-dish is a powerful in vitro platform to study cell-cell interactions in MASH pathogenesis and identify itaconate crosstalk between Kupffer cells and hepatocytes as a novel pathogenic mechanism and a potential therapeutic target in MASH.

biochemistry↗

bioRxiv2026

Hepatic stellate cells drive hypoxia-inducible factor (HIF)-dependent metabolic rewiring and proliferation in hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer in adults and represents the third most common cause of cancer-related deaths worldwide. With limited treatment options, HCC remains a major global health problem. The majority of HCC develops in a cirrhotic liver, where the tumor microenvironment (TME) is enriched in fibroblasts derived from activated hepatic stellate cells (HSCs). Cell-cell interactions in TME play an important role in metabolic reprogramming, which is increasingly recognized as a crucial driver in HCC proliferation. The tumor-promoting and metabolic effects of HSCs on HCC cells however remain incompletely characterized. Understanding how HSC-HCC interaction regulates HCC proliferation can lead to identification of novel therapeutic targets. Here, we established a Transwell co-culture model of HSCs and HCC cells to investigate how HSC-HCC interactions regulate HCC metabolism and proliferation. Using isotope tracing metabolomics, we show that co-culture with HSCs increases proliferation of HCC cells by rewiring glucose metabolism to increase glycolysis and de novo synthesis of nucleotides. Transcriptomic and immunofluorescence staining analyses revealed that HSCs upregulate hypoxia-inducible factor (HIF) signaling in HCC cells, which contributes to metabolic rewiring and HCC proliferation. We further identified H2O2 as a candidate crosstalk signal that could mediate HSC-dependent activation of HIF signaling in HCC cells. Together, these findings reveal a previously overlooked modulatory role of HSC-HCC interaction in metabolism-driven HCC progression. Targeting these pathways may represent a promising strategy to improve HCC treatment.

cancer biology↗

bioRxiv2026

Full Scan enhanced Dynamic Range MS improves metabolite coverage and cancer cell-line discrimination in untargeted metabolomics

Metabolite detection with mass spectrometry (MS) in untargeted metabolomics is limited by the wide concentration range of metabolites, where high-abundance signals dominate MS1 scans and suppress detection of low-abundance features. This reduces metabolite coverage and obscures biologically relevant signals, particularly in complex cellular systems. Full Scan enhanced Dynamic Range (eDR) MS addresses these limitations by partitioning the MS1 mass range into multiple subscans and mass windows, reducing saturation effects from dominant ions. Here, we systematically evaluate different eDR acquisition strategies for untargeted metabolomics. Across four hepatocellular carcinoma cell lines, Full Scan eDR MS increased detectable features up to [~]3.5-fold compared to Full Scan MS. Among equidistant window configurations, 12 windows yielded the highest feature count and broadest dynamic range, while custom window distributions further improved detection in ion-dense regions. In particular, allocating smaller window sizes to the low m/z region selectively increased detection of low-mass features while preserving performance for higher mass ions. Full Scan eDR MS also improved data quality, reducing variation and increasing signal-to-noise ratios, especially for low-abundance metabolites. MS2 coverage and metabolite identifications increased substantially, resulting in unique detection of cancer-relevant metabolites. Importantly, the increased depth of metabolite detection enabled improved discrimination between cancer cell lines, supporting deeper interrogation of metabolic heterogeneity. Overall, these results establish Full Scan eDR MS as a flexible strategy to improve sensitivity and metabolome coverage in untargeted metabolomics. Customization of window size and distribution enable targeted expansion of dynamic range within predefined mass regions, allowing MS acquisition to be tailored to sample complexity and metabolites of interest.

biochemistry↗

bioRxiv2026

hENT Inhibition Prevents Pyrimidine-Driven Resistance to DHODH Inhibition in Malignant Rhabdoid Tumors

Extracranial malignant rhabdoid tumors (ecMRTs) are aggressive pediatric cancers characterized by mutations in genes encoding members of the SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex, with limited effective treatment options. Metabolic reprogramming, including nucleotide biosynthesis, is a hallmark of cancer that represents a potential therapeutic vulnerability. Previously, we demonstrated that inhibition of the de novo pyrimidine synthesis enzyme dihydroorotate dehydrogenase (DHODH) represents a promising treatment strategy for rhabdoid tumors, including ecMRTs and atypical teratoid/rhabdoid tumors (AT/RTs). Using patient-derived tumoroid models, we now extend these findings by showing that another SWI/SNF-deficient pediatric cancer, small cell carcinoma of the ovary, hypercalcemic type (SCCOHT), is also sensitive to DHODH inhibition. Gene expression analyses further confirmed that SCCOHT and AT/RT, like ecMRT, exhibit elevated expression of de novo nucleotide synthesis genes. To determine whether metabolic features of the tumor microenvironment (TME) influence DHODH inhibitor (DHODHi) response, we profiled plasma and tumor interstitial fluid from orthotopic ecMRT-bearing mice and found that the TME was markedly enriched in nucleosides and nucleobases. Supplementation of standard culture media with these nucleosides demonstrated that pyrimidines, but not purines, can rescue ecMRT cells from DHODHi-induced growth inhibition via activation of the pyrimidine salvage pathway. This resistance mechanism was effectively reversed by co-treatment with human equilibrative nucleoside transporter (hENT) inhibitors, enhancing DHODHi sensitivity in ecMRTs. Collectively, these findings reveal a conserved metabolic vulnerability across SWI/SNF-deficient pediatric cancers and emphasize the critical importance of modeling tumor metabolism under physiologically relevant conditions to accurately identify metabolism-targeted therapeutic strategies for these currently lethal pediatric cancer types.

cancer biology↗

bioRxiv2025

Identification of picornavirus proteins that inhibit de novo nucleotide synthesis during infection

The picornavirus family includes well-known pathogens for humans and animals, such as enteroviruses (e.g. poliovirus, coxsackievirus, rhinovirus) and cardioviruses (e.g. encephalomyocarditis virus [EMCV] and Saffold virus). Picornaviruses modulate cellular metabolism likely to generate sufficient building blocks for virus replication. Previously, we showed that coxsackievirus B3 (CVB3) and EMCV remodel nucleotide metabolism during infection. Here, we investigated whether this modulation is attributable to specific viral proteins. For this, we studied the modulation of metabolism by several recombinant CVB3 and EMCV viruses in HeLa cells. Using isotope tracing metabolomics with three distinct labels, 13C6-glucose or 13C5/15N2-glutamine, we reveal that the 2A protease of CVB3 and the Leader protein of EMCV inhibit de novo nucleotide synthesis. Furthermore, we show that nucleotide metabolism is also reprogrammed by CVB3 and EMCV in human induced pluripotent stem cell-derived cardiomyocytes. Our insights are important to increase understanding of picornavirus-host interactions and may lead to novel therapeutic strategies.

microbiology↗

bioRxiv2023

AMPK activation induces RALDHhigh tolerogenic dendritic cells through rewiring of glucose and lipid metabolism

It is well known that dendritic cell (DC) activation and function are underpinned by profound changes in cellular metabolism. Several studies indicate that the ability of DCs to promote tolerance is dependent on catabolic metabolism. The AMP-activated kinase (AMPK) is a central nutrient and energy sensor whose activation promotes catabolism while inhibiting ATP-consuming anabolic pathways. Yet the contribution of AMPK activation to DC tolerogenicity remains unknown. Here, we show that AMPK activation renders human monocyte-derived DCs tolerogenic as evidenced by an enhanced ability to drive differentiation of regulatory T cells, a process dependent on increased RALDH activity. This is accompanied by a number of distinct metabolic changes, in particular increased breakdown of glycerophospholipids, enhanced mitochondrial fission-dependent fatty acid oxidation, and upregulated glucose catabolism. This metabolic rewiring is functionally important as we found interference with these metabolic processes to reduce to various degrees AMPK-induced RALDH activity as well as the tolerogenic capacity of moDCs. Altogether, our findings reveal a key role for AMPK signaling in shaping DC tolerogenicity, and suggest that AMPK may serve as new target to direct DC-driven immune responses in therapeutic settings.

immunology↗

bioRxiv2022

TNFR2 costimulation differentially impacts regulatory and conventional CD4+ T-cell metabolism

CD4+ conventional T cells (Tconvs) mediate adaptive immune responses, whereas regulatory T cells (Tregs) suppress those responses to safeguard the body from autoimmunity and inflammatory diseases. The opposing activities of Tconvs and Tregs depend on the stage of the immune response and their environment, with an orchestrating role for cytokine- and costimulatory receptors. Nutrient availability also impacts T-cell functionality via metabolic and biosynthetic processes that are largely unexplored. Many data argue that costimulation by Tumor Necrosis Factor Receptor 2 (TNFR2) favors support of Treg over Tconv responses and therefore TNFR2 is a key clinical target. Here, we review the pertinent literature on this topic and highlight the newly identified role of TNFR2 as a metabolic regulator for thymus-derived (t)Tregs. We present novel transcriptomic and metabolomic data that show the differential impact of TNFR2 on Tconv and tTreg gene expression and reveal distinct metabolic impact on both cell types.

immunology↗

bioRxiv2020

The lysosomal V-ATPase B1 subunit in proximal tubule is linked to nephropathic cystinosis

Variants in ATP6V1B1, the gene encoding the B1 subunit of the vacuolar H+-ATPase lead to distal renal tubular acidosis and hearing loss of variable degree. Apart from metabolic acidosis, a 23-month-old girl with pathogenic ATP6V1B1 variants presented with increased granulocyte cystine levels, and symptoms of renal Fanconi syndrome, indicating an unknown link to proximal tubulopathy and cystinosis, a disorder of the proximal tubule caused by variants in the CTNS gene. Here, we demonstrated that ATP6V1B1 is expressed in proximal tubules of human kidney tissue, though to a lesser extent than in distal tubules. Further, we used CRISPR/Cas9 technology to selectively knockout ATP6V1B1 or CTNS in human renal proximal tubule cells and performed a full metabolomic and proteomic analysis to compare their phenotype to isogenic control cells. In line with the clinical data, the loss of ATP6V1B1 disrupted the chemiosmotic coupling between the lysosomal V ATPase B1 subunit and cystinosin and resulted in intralysosomal accumulation of cystine and autophagy activation in renal proximal tubule cells. In conclusion, we identified ATP6V1B1 as a central player in renal proximal tubule cells, regulating lysosomal cystine transport and autophagy, and its absence can lead to proximal tubule dysfunction.

pharmacology and toxicology↗

bioRxiv2020

Cysteamine-bicalutamide combination treatment restores alpha-ketoglutarate and corrects proximal tubule phenotype in cystinosis

Nephropathic cystinosis is a severe monogenetic kidney disorder caused by mutations in CTNS, encoding the lysosomal transporter cystinosin, resulting in lysosomal cystine accumulation. The sole treatment, cysteamine, slows down the disease progression, but does not correct the established proximal tubulopathy. Here, we developed a new therapeutic strategy by applying an omics-based strategy to expand our knowledge on the complexity of the disease and prioritize drug targets in cystinosis. We identified alpha-ketoglutarate as a key metabolite linking cystinosin loss, lysosomal autophagy defect and proximal tubular impairment in cystinosis. This insight offered a bicalutamide-cysteamine combination treatment as a novel dual target pharmacological approach for the phenotypical correction of cystinotic proximal tubule cells, patient-derived kidney tubuloids and cystinotic zebrafish.

pharmacology and toxicology↗

bioRxiv2018

Refolding activity of bacterial Hsp90 in vivo reveals ancient chaperoning function

The conserved molecular chaperones Hsp70 and Hsp90 play a key role in protein folding. Mechanistically, Hsp90 acts downstream from Hsp70 solving an Hsp70-inflicted folding block. It is unclear, though, when and to which extend the concerted action of this cascade becomes crucial in living organisms. Here we show that, in E. coli cells, Hsp90 dramatically improves protein refolding after heat stress while it is dispensable for de novo folding. We found that Hsp90 inhibition effectively reduced the refolding yields in vivo, leading to strongly reduced enzymatic activity of the paradigmatic chaperone client luciferase and broadly increased aggregation of the E. coli proteome. Additionally, the presence of Hsp90 during refolding reduces the net ATP consumption presumably by sparing the substrate binding-and-release cycles on Hsp70. This mechanism explains how the cooperation of Hsp90 with the Hsp70 chaperone system creates robust folding machinery in a sustainable manner. Together, we describe a general function for bacterial Hsp90 as a key factor of the folding cascade, which may be the ancient activity of this evolutionary conserved machine.

biochemistry↗