Biology subjects

Broenstrup, M.

Publications and source records attributed to Broenstrup, M..

6 recordsLinked to original sources

bioRxiv2025

Itaconate is metabolized to 2-hydroxymethylsuccinate through a CoA-independent degradation pathway in mitochondria

The immunometabolite itaconate modulates cellular metabolism and is converted into structurally similar C5 dicarboxylates that require advanced analytics to decipher their metabolic fate. Here, we employ high-resolution mass spectrometry and tracing approaches and identify 2-hydroxymethylsuccinate (2HMS) as a previously unrecognized C5 dicarboxylate derived from itaconate. 2HMS synthesis occurs during inflammatory responses and upon itaconate treatment, as detected by 13C itaconate tracing. Pathway analysis reveals that methylglutaconyl-CoA hydratase (AUH) drives 2HMS synthesis through a CoA-independent conversion (CIC) pathway. This pathway is distinct from the CoA-dependent conversion (CDC) pathway that generates mesaconate and itaconyl-CoA influencing B12-dependent processes. In vivo inflammation studies reveal that adipose tissue prefers CIC to produce 2HMS and liver favors CDC-mediated mesaconate synthesis, highlighting tissue-specific itaconate degradation routes. This study identifies a new branch of itaconate metabolism, provides an analytical framework to resolve C5 dicarboxylate networks, and links 2HMS to inflammation and mitochondrial metabolism that might be targeted therapeutically.

cell biology↗

bioRxiv2025

Metabolic reprogramming of methylthioadenosine-dependent sulfur recycling is a major driver of CHIKV infection

The supply of key metabolites into viral replication compartments must be assured through a coordinated reprogramming of host metabolic pathways. For chikungunya virus (CHIKV), the cellular metabolites required for a successful infection are largely unknown. We show that CHIKV reprograms sulfur-dependent pathways. To maintain the resupply of thiols, the methionine (Met) salvage players, 5'-methylthioadenosine (MTA) and methionine adenosyltransferase-2a (Mat2a) are co-induced specifically. Under sulfur-depleted conditions, exogenously added MTA restores CHIKV replication more efficiently than its precursor S-adenosylmethionine, while inhibitions of Mat2a or de novo cysteine (Cys) biosynthesis reduce viral infectivity. Upon sulfur insufficiency CHIKV upregulates the U34-tRNA methyltransferase ALKBH8, and when ALKBH8 is deleted, virus replication is reduced by impairing sulfur relay, recapitulating Met-Cys deprivation effects. We found that MTA-mediated CHIKV replication occurs via m6A-independent priming, and that the S-adenosylhomocysteine hydrolase inhibitors DzNep and Adox, inhibited CHIKV replication with nanomolar potency. Our findings uncovered a major pro-CHIKV metabolic rheostat regulating tRNA modifications that can be targeted with host-directed antiviral agents.

microbiology↗

bioRxiv2024

Predicting antimicrobial class specificity of small molecules using machine learning

Whilst the useful armory of antibiotic drugs is continually depleted due to the emergence of drug-resistant pathogens, the development of novel therapeutics has also slowed down. In the era of advanced computational methods, approaches like machine learning (ML) could be one potential solution to help reduce the high costs and complexity of antibiotic drug discovery and attract collaboration across organizations. In our work, we developed a large antimicrobial knowledge graph (AntiMicrobial-KG) as a repository for collecting and visualizing public in-vitro antibacterial assay. Utilizing this data, we build ML models to efficiently scan compound libraries to identify compounds with the potential to exhibit antimicrobial activity. Our strategy involved training seven classic ML models across six compound fingerprint representations, of which the Random Forest trained on the MHFP6 fingerprint outperformed, demonstrating an accuracy of 75.9% and Cohens Kappa score of 0.68. Finally, we illustrated the models applicability for predicting the antimicrobial properties of two small molecule screening libraries. Firstly, the EU-OpenScreen library was tested against a panel of Gram-positive, Gram-negative, and Fungal pathogens. Here, we unveiled that the model was able to correctly predict more than 30% of active compounds for Gram-positive, Gram-negative, and Fungal pathogens. Secondly, with the Enamine library, a commercially available HTS compound collection with claimed antibacterial properties, we predicted its antimicrobial activity and pathogen class specificity. These results may provide a means for accelerating research in AMR drug discovery efforts by carefully filtering out compounds from commercial libraries with lower chances of being active.

bioinformatics↗

bioRxiv2024

Single influenza A viruses induce nanoscale cellular reprogramming at the virus-cell interface

Viruses, as nanoscale entities with limited proteomes, must efficiently infect target cells using their available resources. During infection, individual virions induce specific cellular signaling within the virus-cell interface, a nanoscale patch of the plasma membrane in contact with the virus. However, virus-induced receptor recruitment and cellular activation are transient processes that occur within minutes and at the nanoscale level. Hence, the temporal and spatial kinetics of such early events often remain poorly understood due to technical limitations. To address this challenge, we developed a novel protocol to covalently immobilize unmodified influenza A viruses on glass surfaces before exposing them to live epithelial cells. This approach extends the observation time for virus-plasma membrane interaction while preserving the viruses native state for uncompromised cell interaction. Using single-molecule super-resolution microscopy, we investigated virus-receptor interaction showing that viral receptors are not immobilized by the virus but rather slowed down, which leads to a specific local receptor accumulation and turnover. We further followed the dynamics of clathrin-mediated endocytosis at the single-virus level and demonstrate the recruitment of adaptor protein 2 (AP-2), previously thought to be uninvolved in influenza A virus infection. Finally, we examined the nanoscale organization of the actin cytoskeleton at the virus-binding site, showing a local and dynamic response of the cellular actin cortex to the infecting virus. Our findings provide novel insights into the fundamental process of virus-cell interaction and demonstrate the versatility and potential impact of our approach on virus-cell biology.

microbiology↗

bioRxiv2023

Rapid enzymatic detection of Shigatoxin-producing E. coli using fluorescence-labeled oligonucleotide substrates

Shigatoxin-producing E. coli (STEC) are important human pathogens causing disease ranging from diarrhea to severe hemolytic uremic syndrome. As STEC are transmitted via animals, food, and water, and may produce large outbreaks, their timely and qualified detection including isolate recovery is of high importance, but challenging and labor-intense. Thus, the availability of an easy-to-perform rapid test would be a tremendous advance. Since the common feature and major virulence factor of otherwise multifaceted STEC is the Shiga toxin (Stx), we developed a detection method for Stx, specifically for its catalytic RNA-N-glycosidase activity targeting the Sarcin Ricin Loop (SRL) of 28S ribosomal RNA. To this end, synthetic ssDNA substrates mimicking the SRL were designed and linked to a fluorophore and quencher pair, which conferred a fluorescence signal after cleavage by Stx. Optimal results using bacterial culture supernatants or single colonies were achieved for substrate StxSense 4 following 30 to 60 minutes incubation. Importantly, different Stx1 and Stx2 subtypes, diverse STEC serotypes, and Shigella were detected. In conclusion, the assay offers rapid and facile detection of STEC based on a real-time readout for Stx activity. Therefore, it may improve STEC risk evaluation, therapy decisions, outbreak and source detection, and simplify research for antimicrobials.

microbiology↗

bioRxiv2021

Repurposing screen highlights broad-spectrum coronavirus antivirals and their host targets

Libraries composed of licensed drugs represent a vast repertoire of molecules modulating physiologic processes in humans, thus providing unique opportunities for discovery of host targeting antivirals. We interrogated the ReFRAME repurposing library with 12,993 molecules for broad-spectrum coronavirus antivirals and discovered 134 compounds inhibiting an alphacoronavirus, mapping to 59 molecular target categories. Dominant targets included the 5-hydroxytryptamine receptor and dopamine receptor and cyclin-dependent kinase inhibitors. Counter-screening with SARS-CoV-2 and validation in primary cells identified Phortress, an aryl hydrocarbon receptor (AHR) ligand, Bardoxolone and Omaveloxolone, two nuclear factor, erythroid 2 like 2 (NFE2L2) activators as inhibitors of both alpha- and betacoronaviruses. The landscape of coronavirus targeting molecules provides important information for the development of broad-spectrum antivirals reinforcing pandemic preparedness.

microbiology↗