Biology subjects

Skruber, K.

Publications and source records attributed to Skruber, K..

10 recordsLinked to original sources

bioRxiv2026

Hailey-Hailey disease models identify synergistic therapeutic effects of MEK and ROCK inhibition

Hailey-Hailey disease (HHD) is a genetic skin blistering disorder lacking approved treatments despite linkage to ATP2C1 variants 25 years ago. Since knockout mice did not replicate HHD, we ablated ATP2C1 in human keratinocytes or chemically inhibited its encoded Golgi calcium pump SPCA1. In organotypic epidermis, SPCA1 deficiency or inhibition reproduced HHD pathology, disrupting desmosomal cadherins and severing cell-cell junctions, termed acantholysis. RNA sequencing of heterozygous cells identified dysregulation of actin and Rho GTPases along with EGF receptor signaling as potential pathogenic drivers. Accordingly, SPCA1-depleted organotypic epidermis and HHD biopsies exhibited cortical actin disorganization and hyper-phosphorylation of the Rho kinase (ROCK) target, myosin light chain. Rho activation was sufficient to induce acantholysis, while ROCK inhibition partially restored heterozygous keratinocyte cohesion. A fluorescent biosensor demonstrated ERK hyper-activation in heterozygous cells along with desmosomal cadherin mis-localization. Importantly, treating SPCA1-deficient keratinocyte sheets with MEK and ROCK inhibitors together fully restored their integrity. Our results show HHD blistering is driven by desmosome and cortical actin dysfunction that was mitigated by targeting MEK and ROCK with repurposed drugs, offering a viable treatment strategy. Moreover, our model provides a blueprint for replicating genetic epidermal disorders to delineate pathogenic mechanisms and vet therapeutics for other orphan skin diseases. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/726679v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@b2de6aorg.highwire.dtl.DTLVardef@128411borg.highwire.dtl.DTLVardef@1ca760forg.highwire.dtl.DTLVardef@10cd6c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

A geothermal amoeba sets a new upper temperature limit for eukaryotes

The study of temperature limits has transformed our knowledge of the boundaries of life but has been largely focused on bacteria and archaea. We isolated a novel geothermal amoeba, Incendiamoeba cascadensis, that divides at 63{degrees}C (145.4{degrees}F), establishing a new record for the upper temperature limit across all eukaryotes. We demonstrated cellular proliferation with growth experiments and visualized mitosis via expansion microscopy. Using high-temperature live-cell imaging, we quantified movement up to 64{degrees}C. We assembled the genome of I. cascadensis and using comparative genomics found an enrichment of genes related to proteostasis, genome stability, and sensing the external environment. Taken together, our findings challenge the current paradigm of temperature constraints on eukaryotic cells and reshape our understanding of where and how eukaryotic life can persist.

microbiology↗

bioRxiv2025

Membrane-associated polymerases deliver most of the actin subunits to a lamellipodial network

Actin filaments are two-stranded protein polymers that form the basic structural unit of the eukaryotic actin cytoskeleton. While filaments assembled from purified actin in vitro elongate when soluble monomers bind to free filament ends, in cells the mechanism of filament elongation is less clear. Most monomeric actin in the cytoplasm is bound to the accessory protein profilin, and many regulators of filament assembly recruit actin-profilin complexes to membrane surfaces where they locally accelerate filament elongation. Employing quantitative live-cell imaging of actin-profilin fusion proteins and biochemically defined mutants of the branched actin regulator, WAVE1, we find that only [~]25% of the actin in leading-edge lamellipodial networks enters directly from solution, while the majority enters via membrane-associated polymerases.

cell biology↗

bioRxiv2024

Layilin Regulates Treg Motility and Suppressive Capacity in Skin

Regulatory T cells (Tregs) are essential for maintaining immune tolerance in both lymphoid and non-lymphoid tissues. We discovered that layilin, a C-type lectin receptor, is predominantly expressed on Tregs in skin. Layilin was highly expressed on a subset of clonally expanded effector Tregs in both healthy and psoriatic skin. Layilin expressing Tregs exhibited a transcriptional profile indicative of enhanced adhesion. Deletion of layilin in Tregs in vivo resulted in significantly attenuated skin inflammation. Mechanistically, layilin enhanced Treg adhesion via modulation of LFA-1, resulting in distinct cytoskeletal alterations consistent with enhanced focal adhesion and lamellipodia formation. Taken together, we define layilin as a critical regulator of Treg suppressive capacity through modulating motility and adhesion in a non-lymphoid tissue.

immunology↗

bioRxiv2024

Actin filament assembly driven by distributive polymerases clustered on membrane surfaces

Actin filaments created by the Arp2/3 complex form branched networks, that grow and push against cellular membranes. We employ theory and simulation to describe how membrane surfaces accelerate filament assembly via clustering of proteins that bind actin monomers and/or profilin-actin complexes. Briefly, thermal fluctuations drive filament tips on constrained, two-dimensional random walks across the membrane, where they encounter multiple actin-charged polymerases. At low actin concentrations, filament elongation is limited by delivery of monomers to the membrane surface; at high actin concentrations, elongation depends on how quickly fluctuating filaments search the membrane. Using experimentally measured parameter values we conclude that surface-mediated polymerization can outpace solution-mediated elongation, even at high actin concentrations (>200 {micro}M). The finite time required for profilin dissociation decreases the advantage conferred by surface-associated polymerases, but only in the absence of force. Load forces enhance the effect of surface polymerases, which can both accelerate elongation and increase the force required to stall filament assembly.

cell biology↗

bioRxiv2023

Control of microtubules in neuronal processes by profilin 1 and actomyosin

Microtubules, intermediate filaments, and actin are cytoskeletal polymer networks found within the cell. While each has unique functions, all the cytoskeletal elements must work together for cellular mechanics to be fully operative. This is achieved through crosstalk mechanisms whereby the different networks influence each other through signaling pathways and direct interactions. Because crosstalk can be complex, it is possible for perturbations in one cytoskeletal element to affect the others in ways that are difficult to predict. Here we investigated how long-term changes to the actin cytoskeleton affect microtubules and intermediate filaments. Reducing F-actin or actomyosin contractility increased acetylated microtubules and intermediate filament expression, with the effect being significantly more pronounced in neuronal processes. Changes to microtubules were completely reversible if F-actin and myosin activity is restored. Moreover, the altered microtubules in neuronal processes resulting from F-actin depletion caused significant changes to microtubule-based transport, mimicking phenotypes that are linked to neurodegenerative disease. Thus, defects in actin dynamics cause a compensatory response in other cytoskeleton components which profoundly alters cellular function.

cell biology↗

bioRxiv2023

The actin binding protein profilin 1 is critical for mitochondria function

Profilin 1 (PFN1) is an actin binding protein that is vital for the polymerization of monomeric actin into filaments. Here we screened knockout cells for novel functions of PFN1 and discovered that mitophagy, a type of selective autophagy that removes defective or damaged mitochondria from the cell, was significantly upregulated in the absence of PFN1. Despite successful autophagosome formation and fusion with the lysosome, and activation of additional mitochondrial quality control pathways, PFN1 knockout cells still accumulate damaged, dysfunctional mitochondria. Subsequent imaging and functional assays showed that loss of PFN1 significantly affects mitochondria morphology, dynamics, and respiration. Further experiments revealed that PFN1 is located to the mitochondria matrix and is likely regulating mitochondria function from within rather than through polymerizing actin at the mitochondria surface. Finally, PFN1 mutants associated with amyotrophic lateral sclerosis (ALS) fail to rescue PFN1 knockout mitochondrial phenotypes and form aggregates within mitochondria, further perturbing them. Together, these results suggest a novel function for PFN1 in regulating mitochondria and identify a potential pathogenic mechanism of ALS-linked PFN1 variants.

cell biology↗

bioRxiv2021

TDAExplore: quantitative image analysis through topology-based machine learning

Machine learning has greatly expanded the ability to classify images. However, many machine learning classifiers require thousands of images for training and lack quantitative descriptors of how images were grouped. We overcome these limitations with a machine learning approach based on topological data analysis, where a data set of 20-30 images is sufficient to accurately train the classifier. Our method quantifies differences between groups and identifies subcellular regions with the largest dissimilarities.

cell biology↗

bioRxiv2019

Profilin 1 Controls the Assembly, Organization, and Dynamics of Leading Edge Actin Structures Through Internetwork Competition and Collaboration

How actin monomers are distributed to different networks remains poorly understood. One emerging concept is that the monomer pool is limited and heterogenous, causing biased assembly and internetwork competition. However, most knowledge regarding monomer distribution comes from studies where competing networks are discrete. In metazoans, many actin-based structures are complex, containing competing networks that overlap and are functionally interdependent. Addressing how monomers control the assembly and organization of these complex structures is critical to understanding how actin functions in cells. Here, we identify the monomer-binding protein profilin 1 (PFN1) as a major determinant of actin assembly, organization, and network homeostasis in mammalian cells. At the leading edge, PFN1 controls the localization and activity of the assembly factors Arp2/3 and Mena/VASP, with discrete stages of internetwork competition and collaboration occurring at different PFN1 concentrations. This causes substantial changes to leading edge actin architecture and the types of structures that form there.

cell biology↗

bioRxiv2018

Enhanced Neurite Outgrowth and Regeneration in ALS Resistant Motor Neurons from SOD1 Mutant Mouse Models

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal neurodegenerative disease characterized by motor neuron cell death. However, not all motor neurons are equally susceptible. Most of what we know about the surviving motor neurons comes from gene expression profiling, less is known about their functional traits. We found that resistant motor neurons cultured from SOD1 ALS mouse models have enhanced axonal outgrowth and dendritic branching. They also have an increase in the number and size of actin-based structures like growth cones and filopodia. These phenotypes occur in cells cultured from presymptomatic mice and mutant SOD1 models that do not develop ALS, but not in embryonic motor neurons. Enhanced outgrowth and upregulation of filopodia can be induced in wild-type adult cells by expressing mutant SOD1. These results demonstrate that mutant SOD1 can enhance the regenerative capability of ALS resistant motor neurons. Capitalizing on this mechanism could lead to new therapeutic strategies.

neuroscience↗