Biology subjects

Metzner, K.

Publications and source records attributed to Metzner, K..

6 recordsLinked to original sources

bioRxiv2026

Micafungin exposure drives multidrug resistance in Clavispora lusitaniae

Fungal infections are an escalating global health concern, with rare Candida species posing an urgent threat due to emerging multidrug resistance. Clavispora (Candida) lusitaniae is an uncommon pathogen in which multidrug resistance has been documented during antifungal therapy, yet the selective forces driving this phenotype remain unclear. Here, we show that exposure to the echinocandin micafungin (MCF) alone can select for multidrug resistance in C. lusitaniae. Through controlled evolution experiments we identified individual point mutations in genes encoding ergosterol biosynthesis enzymes (ERGs), sterol trafficking proteins (OSH2), and the echinocandin drug target (FKS1) that confer a significant fitness benefit to one or more classes of antifungals. We find that ERG3 loss-of-function is the primary and independent driver of pan-antifungal resistance to echinocandins, azoles and polyenes. The ERG3 mutants have <1% ergosterol, increased levels of non-toxic sterol intermediates, and increased chitin content, consistent with both cell membrane and cell wall remodeling that enables the fungal pathogen to evade all three drug classes. The convergence of sterol reprogramming and compensatory cell wall remodeling that occurs during adaptation to echinocandin monotherapy can evolve through a single point mutation and parallels our recent case study of acquired multidrug resistance. IMPORTANCEMultidrug resistance in Candida species severely limits treatment options and increases mortality, particularly in immunocompromised patients. Despite increasing reports of multidrug resistance, the molecular mechanisms driving multidrug resistance remain poorly understood. We find that in vitro MCF exposure alone can drive multidrug resistance in C. lusitaniae via acquisition of de novo point mutations in ERG3, an observation that parallels our recent patient case study. By identifying causative mutations and associated physiological changes, we provide mechanistic insight into the emergence of multidrug resistance and highlight the need for surveillance strategies that account for resistance evolution under echinocandin monotherapy.

microbiology↗

bioRxiv2024

Comparative Evaluation of Bioinformatic Pipelines for Full-Length Viral Genome Assembly

The increasingly widespread application of next-generation sequencing (NGS) in clinical diagnostics and epidemiological research has generated a demand for robust, fast, automated, and user-friendly bioinformatic workflows. To guide the choice of tools for the assembly of full-length viral genomes from NGS datasets, we assessed the performance and applicability of four widely adopted bioinformatic pipelines (shiver - for which we created a user-friendly Dockerized version, referred to as dshiver; SmaltAlign, viral-ngs, and V-pipe) using both simulated datasets and real-world HIV-1 paired-end short- read sequences and default settings. All four pipelines produced high-quality consensus genome assemblies and minority variant calls when the reference sequence used for assembly had high similarity to the analyzed sample. However, while shiver and SmaltAlign showed robust performance also with more divergent samples (non-matching subtypes), viral-ngs and V-Pipe proved to be sensitive to genetic distance from the reference sequence. With empirical datasets, SmaltAlign and viral-ngs exhibited substantially shorter runtime compared to V-Pipe and shiver. In terms of applicability, V-Pipe provides the broadest functionalities; SmaltAlign and dshiver combine user-friendliness with robustness; while the use of viral-ngs requires a less computational resources compared to other tools. To conclude, all four pipelines can perform well in terms of quality metrics; however, the reference sequence needs to be adjusted to closely match the sample data for viral-ngs and V-Pipe. Differences in user-friendliness and runtime may guide the choice of the pipeline in a particular setting. The new Dockerized version of shiver offers ease of use in addition to the accuracy and robustness of the original pipeline.

bioinformatics↗

bioRxiv2024

Step-wise evolution of azole resistance through copy number variation followed by KSR1 loss of heterozygosity in Candida albicans.

Antimicrobial drug resistance poses a global health threat, requiring a deeper understanding of the evolutionary processes that lead to its emergence in pathogens. Complex evolutionary dynamics involve multiple mutations that can result in cooperative or competitive (clonal interference) effects. Candida albicans, a major fungal pathogen, displays high rates of copy number variation (CNV) and loss of heterozygosity (LOH). CNV and LOH events involve large numbers of genes and could synergize during evolutionary adaptation. Understanding the contributions of CNV and LOH to antifungal drug adaptation is challenging, especially in the context of whole-population genome sequencing. Here, we document the sequential evolution of fluconazole tolerance and then resistance in a C. albicans isolate involving an initial CNV on chromosome 4, followed by an LOH on chromosome R that involves KSR1. Similar LOH events involving KSR1, which encodes a reductase involved in sphingolipid biosynthesis, were also detected in independently evolved fluconazole resistant isolates. We dissect the specific KSR1 codons that affect fluconazole resistance and tolerance. The combination of the chromosome 4 CNV and KSR1 LOH results in a >500-fold increase in azole resistance, illustrating a compelling example of rapid, yet step-wise, interplay between CNV and LOH in drug resistance evolution.

microbiology↗

bioRxiv2023

Discovery of a new activator of Slack potassium channels with robust efficacy in models of histamine-independent and chronic itch

Various disorders are accompanied by histamine-independent itching, which is often resistant to the currently available therapies. In this study, we hypothesized that pharmacological activation of Slack (Kcnt1, KNa1.1), a potassium channel highly expressed in itch-sensitive sensory neurons, has therapeutic potential for the treatment of itching. Based on the Slack-activating antipsychotic drug, loxapine, we designed a series of new derivatives with improved pharmacodynamic and pharmacokinetic profiles that enabled us to validate Slack as a pharmacological target in vivo. One of these new Slack activators, compound 6, exhibited negligible dopamine D2 and D3 receptor binding, unlike loxapine. We found that compound 6 displayed potent on-target antipruritic activity in multiple mouse models of acute histamine-independent and chronic itch without motor side effects. These properties make compound 6 a lead molecule for the development of new antipruritic therapies targeting Slack.

pharmacology and toxicology↗

bioRxiv2023

Imaging-based screening identifies modulators of the eIF3 translation initiation factor complex in Candida albicans

Fungal pathogens like Candida albicans can cause devastating human disease. Treatment of candidemia is complicated by the high rate of resistance to common antifungal therapies. Additionally, there is host toxicity associated with many antifungal compounds due to the conservation between essential mammalian and fungal proteins. An attractive new approach for antimicrobial development is to target virulence factors: non-essential processes that are required for the organism to cause disease in human hosts. This approach expands the potential target space while reducing the selective pressure towards resistance, as these targets are not essential for viability. In C. albicans, a key virulence factor is the ability to transition to hyphal morphology. We developed a high-throughput image analysis pipeline to distinguish between yeast and filamentous growth in C. albicans at the single cell level. Based on this phenotypic assay, we screened the FDA drug repurposing library of 2,017 compounds for their ability to inhibit filamentation and identified 33 compounds that block the hyphal transition in C. albicans with IC50 values ranging from 0.2 to 150 {micro}M. Multiple compounds showed a phenyl vinyl sulfone chemotype, prompting further analysis. Of these phenyl vinyl sulfones, NSC 697923 displayed the most efficacy, and by selecting for resistant mutants, we identified eIF3 as the target of NSC 697923 in C. albicans.

microbiology↗

bioRxiv2021

Molecular properties of Ankrd26 affected by mutations linked to leukemia and carcinoma formation

Derailed signaling originating from the plasma membrane is associated with many types of cancer. Different human cancers and thrombocytopenia are linked to ANKRD26 mutations. We unveil that Ankrd26 is a plasma membrane-localized protein forming nanoclusters and that Ankrd26 is critical for retinoic acid/BDNF-induced neuroblastoma differentiation. An N-terminal amphipathic structure lacking in an AML-associated Ankrd26 mutant is indispensable for membrane binding and bending by partial membrane insertion and renders Ankrd26 inactive in both gain-of-function and loss-of- function/rescue studies addressing cellular differentiation. In a papillary thyroid carcinoma-linked mutant, truncated Ankrd26 is fused with the kinase domain of the protooncogene RET. Our data show that the Ankrd26 part of this fusion mutant mediates anchoring of the RET kinase domain to the plasma membrane and self-association by the coiled coil domain of Ankrd26. Ankrd26-RET fusion led to massively increased ERK1/2 activity and RET autophosphorylation at both Y905 and Y1015, i.e. caused aberrant RET signaling. Our results highlight the importance and molecular details of Ankrd26-mediated organizational platforms for cellular differentiation and signaling pathways from the plasma membrane, which, if derailed, lead to cancer-associated pathomechanisms involving the unveiled Ankrd26 properties.

cancer biology↗