Biology subjects

Wedlich-Soldner, R.

Publications and source records attributed to Wedlich-Soldner, R..

7 recordsLinked to original sources

bioRxiv2025

Effect of very long-chain lipids on the organization of biological membranes: A simulation perspective

Lipids in all biological membranes are distributed heterogeneously across the bilayer. A particularly striking example for this asymmetry is the yeast plasma membrane (PM), which exhibits a high concentration of very long-chain sphingolipids (SL) in its outer leaflet. Experimental observations indicate the existence of highly ordered gel-like PM domains that are enriched in SL but depleted in the major yeast sterol ergosterol. For a better mechanistic understanding of these unusual domains we have performed coarse-grained molecular dynamics simulations with membranes containing varying concentrations of very long-chain lipids. In agreement with experimental results we observed formation of a gel phase, with high order parameter of the acyl chains and hexagonal arrangement of lipid tails, at higher concentrations of very long-chain lipids. Our simulations also show that ergosterol is excluded from these gel regions and that, even when embedded into a liquid disordered phase, gels remain stable on the simulation time scale.

biophysics↗

bioRxiv2024

Structure of the F-tractin-F-actin complex

F-tractin is a short peptide widely used to visualize the actin cytoskeleton in live eukaryotic cells. Similar to other actin-binding probes, F-tractin alters actin organization and impairs cell migration when expressed at high levels. In addition, the probe has been reported to directly induce actin bundling. To elucidate the mechanism behind these effects, we determined the structure of the F-tractin-F-actin complex using electron cryo-microscopy. Our analysis revealed that the F-tractin peptide consists of a flexible N-terminal region and an amphipathic C-terminal helix. The N-terminal part is completely dispensable for F-actin binding but is responsible for the actin bundling effect. The C-terminal helical region interacts with a hydrophobic pocket formed by two neighboring actin subunits, a region identified as an interface for many other actin-binding polypeptides, including Lifeact, the most widely used actin-binding probe. Thus, rather than contrasting F-tractin and Lifeact, our data indicate that these peptides have analogous modes of interaction with F-actin. Our study dissects the structural elements of F-tractin and provides a mechanistic basis for the selection and future development of actin probes.

biochemistry↗

bioRxiv2024

A Red Fluorescent Lifeact Marker to Study Actin Morphology in Podocytes

F-actin is a major component of the cellular cytoskeleton, responsible for maintaining cell shape, enabling movement and facilitating intracellular transport. In the kidney, glomerular podocytes are highly dependent on their actin cytoskeleton shaping their unique foot processes. Hereditary mutations in actin-binding proteins cause focal segmental glomerulosclerosis, while other organs remain largely unaffected. So far, actin visualization in podocytes has been limited to electron microscopy or indirect immunofluorescent labeling of actin-binding proteins. However, the short F-actin-binding peptide Lifeact enables researchers to study actin dynamics in vitro and in vivo with minimal interference with actin metabolism. Here we introduce a new mouse model with conditional expression of a Lifeact.mScarlet-I fusion protein providing red labeling of actin. Cre recombinase-mediated activity allows cell-specific and mosaic expression in podocytes, enabling selective labeling of individual cells to contrast with non-expressing neighboring cells. Transgenic mice are born healthy and young animals display no kidney-related phenotype. By intravital imaging and super-resolution microscopy, we show subcellular localization of actin to the foot processes in a resolution previously only obtainable by electron microscopy. Our novel mouse line provides the opportunity to study the actin cytoskeleton in podocytes and other cell types by intravital imaging and other conventional light microscopy techniques.

cell biology↗

bioRxiv2022

Tetraspanner-based nanodomains modulate BAR domain-induced membrane curvature

Topography is a critical feature driving formation and dynamics of protein and lipid domains within biological membranes. The yeast plasma membrane (PM) has provided a powerful model system to study lateral domain formation, including characteristic BAR domain-induced PM furrows. Currently, it is not clear how the components involved in the establishment of these furrows cooperate to precisely regulate local PM topography. Here we report opposing functions for the Sur7 and Nce102 families of tetraspanner proteins in modulating membrane curvature and domain topography. Using STED nanoscopy and freeze-fracture EM we found that Sur7 tetraspanners form multimeric strands at the upper edges of PM furrows, which counteract the forces exerted by BAR domain proteins and prevent membrane tubulation. In contrast, Nce102 tetraspanners are located basal to the Sur7 proteins and promote BAR domain-induced curvature. The segregation of the two tetraspanner-based nanodomains is further supported by differential distribution of ergosterol to the upper edge of furrows and PIP2 lipids at the furrow base. These findings suggest a general role of tetraspanner proteins in sculpting local membrane domains.

cell biology↗

bioRxiv2019

A deregulated stress response underlies distinct INF2 associated disease profiles

Monogenic diseases provide favorable opportunities to elucidate the molecular mechanisms of disease progression and improve medical diagnostics. However, the complex interplay between genetic and environmental factors in disease etiologies makes it difficult to discern the mechanistic links between different alleles of a single locus and their associated pathophysiologies. Here we systematically characterize a large panel (>50) of autosomal dominant mutants of the actin regulator inverted formin 2 (INF2) that have been reported to cause the podocytic kidney disease focal segmental glomerulosclerosis (FSGS) and the neurological disorder Charcot Marie-Tooth disease (CMT). We found that INF2 mutations lead to deregulated activation of the formin and a constitutive stress response in cultured cells, primary patient cells and Drosophila nephrocytes. Using quantitative live-cell imaging we were able to identify distinct subsets of INF2 variants that exhibit varying degrees of activation. Furthermore, we could clearly distinguish between INF2 mutations that were linked exclusively to FSGS from those that caused a combination of FSGS and CMT. Our results indicate that cellular profiling of disease-associated mutations can contribute substantially to sequence-based phenotype predictions.

cell biology↗

bioRxiv2019

Pervasive head-to-tail insertions of DNA templates mask desired CRISPR/Cas9-mediated genome editing events

CRISPR/Cas9 mediated homology-directed DNA repair is the method of choice for precise gene editing in a wide range of model organisms, including mouse and human. Broad use by the biomedical community refined the method, making it more efficient and sequence specific. Nevertheless, the rapidly evolving technique still contains pitfalls. During the generation of six different conditional knock-out mouse models, we discovered that frequently (sometimes solely) homology-directed repair and/or non-homologous end-joining mechanisms caused multiple unwanted head-to-tail insertions of donor DNA templates. Disturbingly, conventionally applied PCR analysis--in most cases--failed to identify such multiple integration events, which led to a high rate of falsely claimed precisely edited alleles. We caution that comprehensive analysis of modified alleles is essential, and offer practical solutions to correctly identify precisely edited chromosomes.

genetics↗

bioRxiv2018

Lateral plasma membrane compartmentalization links protein function and turnover

Biological membranes organize their proteins and lipids into nano- and microscale patterns. In the yeast plasma membrane (PM) constituents segregate into a large number of distinct domains. However, if and how this intricate patchwork contributes to biological functions at the PM is still poorly understood. Here, we reveal an elaborate interplay between PM compartmentalization, biochemical function and endocytic turnover. Using the methionine permease Mup1 as model system we demonstrate that this transporter segregates into PM clusters. Clustering requires sphingolipids, the tetraspanner Nce102 and TORC2 signaling. Importantly, we show that during substrate transport, a simple conformational change in Mup1 mediates rapid relocation into a unique disperse network at the PM. Clustered Mup1 is protected from turnover, whereas relocated Mup1 actively recruits the endocytic machinery thereby initiating its own turnover. Our findings suggest that lateral compartmentalization provides an important regulatory link between function and turnover of PM proteins.

cell biology↗