Biology subjects

Munk, A.

Publications and source records attributed to Munk, A..

7 recordsLinked to original sources

bioRxiv2026

Distinct mitochondrial DNA single-nucleotide variant signatures in TOP3A-deficient cardiomyocytes

The human heart has a continuous and exceptionally high demand for energy, which is met primarily through mitochondrial oxidative phosphorylation. This dependence places the maintenance and integrity of mitochondrial DNA (mtDNA) for proper mitochondrial function at the center of cardiac health, as mtDNA instability has been shown to cause mitochondrial dysfunction, which can ultimately impair cardiac function leading to cardiomyopathy (CM) and heart failure. However, the contribution of mtDNA instability to cardiac disease remains poorly understood. A growing number of nuclear-encoded proteins have emerged as essential regulators of mtDNA maintenance, organization and segregation. DNA Topoisomerase 3 (TOP3A) is expressed as two isoforms: one is a nuclear-related isoform involved in nuclear genome maintenance while the other localizes to the mitochondria to preserve mtDNA integrity. Recently, individuals bearing biallelic loss-of-function variants in TOP3A manifested phenotypic traits, including CM, typical for mitochondrial dysfunction, supporting a potential mechanistic link between mitochondrial genome instability and TOP3A-related cardiac disease. Here, we investigated the effects of TOP3A deficiency on mtDNA maintenance and stability in the context of TOP3A-associated CM. We employed isogenic wild-type, TOP3A-knockout and BLM-knockout induced pluripotent stem cells (iPSCs) to generate cardiomyocytes (iPSC-CMs) and established a high-throughput, ultra-deep mtDNA sequencing strategy achieving approximately 500,000X coverage to characterize low-frequency mtDNA mutational patterns. Loss of TOP3A triggered an early burst of low-frequency de novo mtDNA single-nucleotide variants during cardiac differentiation, with a striking enrichment within the mitochondrial ribosomal RNA genes, accompanied by a progressive increase in the heteroplasmy of low-frequency mtDNA variants inherited from the common isogenic background. These unique mtDNA signatures were associated with defective mtDNA copy-number expansion and impaired mitochondrial respiration in mature iPSC-CMs. Together, our approach uncovered a previously uncharacterized consequence of TOP3A deficiency and established a link between impaired mtDNA maintenance and mitochondrial dysfunction in TOP3A-associated CM.

genetics↗

bioRxiv2024

Therapeutically targeting the classical complement pathway with antisense oligonucleotides in Alzheimer's disease

The complement classical pathway (CP) is a key mediator of synapse loss and neurodegeneration in mouse models of Alzheimers (AD) and other neurodegenerative diseases. We analyzed human brain proteomics and found consistent elevations of all CP proteins, but not other complement pathways, in AD patient brains. We performed human genetics analysis that identified a rare variant in the C1S gene within the Finnish population that is associated with AD and we found that a common AD-associated C1S variant correlates with increased C1S protein levels. A targeted assay detected elevated C1S activation in AD patient CSF. Given this specific implication of the CP in AD, we next evaluated the therapeutic approach of targeting the CP in the brain using antisense oligonucleotides (ASOs). To identify promising CP targets for knockdown using ASOs we first tested for rescue of synapse loss in an AD mouse model using heterozygous and homozygous complement knockout mice and examined the relative brain expression levels of different CP genes. Based on these experiments we prioritized C1r, C1s and C4 as promising targets for therapeutic knockdown using ASOs. We then screened for ASOs for each target, evaluating in vitro and in vivo knockdown and toxicity, and identified optimal ASOs targeting C1r, C1s and C4. Experiments with AD model mice demonstrated significant rescue of synapse loss following treatment with C1r, C1s or C4 ASOs. Overall, our findings provide proof of concept for using nucleic acid-based medicine to target the CP in AD and demonstrate the translational potential of this approach.

neuroscience↗

bioRxiv2024

MultiMatch: Geometry-Informed Colocalization in Multi-Color Super-Resolution Microscopy

With recent advances in multi-color super-resolution light microscopy it has become possible to simultaneously visualize multiple subunits within complex biological structures at nanometer resolution. To opti-mally evaluate and interpret spatial proximity of stainings on such an image, colocalization analysis tools have to be able to integrate prior knowledge on the local geometry of the recorded biological complex. Here, we present MultiMatch to analyze the abundance and location of chain-like particle arrangements in multi-color microscopy based on multi-marginal optimal unbalanced transport methodology. Our object-based colocalization model statistically addresses the effect of incomplete labeling efficiencies enabling inference on existent, but not fully observ-able particle chains. We showcase that MultiMatch is able to consistently recover all existing chain structures in three-color STED images of DNA origami nanorulers and outperforms established geometry-uninformed triplet colocalization methods in this task in a simulation study. Further-more, MultiMatch also excels in the evaluation of simulated four-color STED images and generalizations to even more color channels can be immediately derived from our analysis. MultiMatch is provided as a user-friendly Python package comprising intuitive colocalization visual-izations and a computationally efficient network flow implementation.

bioinformatics↗

bioRxiv2023

Vimentin intermediate filaments structure and mechanically support microtubules in cells

The eukaryotic cytoskeleton is an intricate network of three types of mechanically distinct biopolymers - actin filaments, microtubules and intermediate filaments (IFs). These filamentous networks determine essential cellular functions and properties. Among them, microtubules are important for intracellular transport and establishing cell polarity during migration. Despite their intrinsic stiffness, they exhibit characteristic bending and buckling in cells due to non thermal forces acting on them. Interactions between cytoskeletal filaments have been found but are complex and diverse with respect to their effect on the mechanical behavior of the filaments and the architecture of networks. We systematically study how actin and vimentin IFs influence the network structure and local bending of microtubules by analyzing fluorescence microscopy images of mouse fibroblasts on protein micropatterns. Our automated analysis averages over large amounts of data to mitigate the effect of the considerable natural variance in biological cell data. We find that the radial orientation of microtubules in circular cells is robust and is established independently of vimentin and actin networks. Observing the local curvature of microtubules, we find highly similar average bending of microtubules in the entire cell regardless of the cytoskeletal surrounding. Small systematic differences cannot be attributed directly to vimentin and actin densities. Our results suggest that, on average, microtubules in unpolarized mouse fibroblasts are unexpectedly independent of the rest of the cytoskeleton in their global network structure and their local response to forces.

biophysics↗

bioRxiv2020

An antibiotic-resistance conferring mutation in a neisserial porin: Structure, ion flux, and ampicillin binding

Gram-negative bacteria cause the majority of highly drug-resistant bacterial infections. To cross the outer membrane of the complex Gram-negative cell envelope, antibiotics permeate through porins, trimeric channel proteins that enable the exchange of small polar molecules. Mutations in porins contribute to the development of drug-resistant phenotypes. In this work, we show that a single point mutation in the porin PorB from Neisseria meningitidis, the causative agent of bacterial meningitis, can strongly affect the binding and permeation of beta-lactam antibiotics. Using X-ray crystallography, high-resolution electrophysiology, atomistic biomolecular simulation, and liposome swelling experiments, we demonstrate differences in drug binding affinity, ion selectivity and drug permeability of PorB. Our work further reveals distinct interactions between the transversal electric field in the porin eyelet and the zwitterionic drugs, which manifest themselves under applied electric fields in electrophysiology and are altered by the mutation. These observations may apply more broadly to drug-porin interactions in other channels. Our results improve the molecular understanding of porin-based drug-resistance in Gram-negative bacteria.

biophysics↗

bioRxiv2019

treeSeg: Testing for Dependence on Tree Structures

Tree structures, showing hierarchical relationships and the latent structures between samples, are ubiquitous in genomic and biomedical sciences. A common question in many studies is whether there is an association between a response variable measured on each sample and the latent group structure represented by some given tree. Currently this is addressed on an ad hoc basis, usually requiring the user to decide on an appropriate number of clusters to prune out of the tree to be tested against the response variable. Here we present a statistical method with statistical guarantees that tests for association between the response variable and a fixed tree structure across all levels of the tree hierarchy with high power, while accounting for the overall false positive error rate. This enhances the robustness and reproducibility of such findings. Significance StatementTree like structures are abundant in the empirical sciences as they can summarize high dimensional data and show latent structure among many samples in a single framework. Prominent examples include phylogenetic trees or hierarchical clustering derived from genetic data. Currently users employ ad hoc methods to test for association between a given tree and a response variable, which reduces reproducibility and robustness. In this paper, we introduce treeSeg, a simple to use and widely applicable methodology with high power for testing between all levels of hierarchy for a given tree and the response while accounting for the overall false positive rate. Our method allows for precise uncertainty quantification and therefore increases interpretability and reproducibility of such studies across many fields of science.

bioinformatics↗

bioRxiv2018

High-resolution experimental and computational electrophysiology reveals weak β-lactam binding events in the porin PorB

The permeation of most antibiotics through the outer membrane of Gram-negative bacteria occurs through porin channels. To design drugs with increased activity against Gram-negative bacteria in the face of the antibiotic resistance crisis, the strict constraints on the physicochemical properties of the permeants imposed by these channels must be better understood. Here we show that a combination of high-resolution electrophysiology, new noise-filtering analysis protocols and atomistic biomolecular simulations reveals weak binding events between the {beta}-lactam antibiotic ampicillin and the porin PorB from the pathogenic bacterium Neisseria meningitidis. In particular, an asymmetry often seen in the electrophysiological characteristics of ligand-bound channels is utilised to characterise the binding site and molecular interactions in detail, based on the principles of electro-osmotic flow through the channel. Our results provide a rationale for the determinants that govern the binding and permeation of zwitterionic antibiotics in anion-selective porin channels.

biophysics↗