- by Zhao, T., Pelegrina-Hidalgo, N., Edwards, D. C., Bak, K. M., Karmakar, U., Fernando, A. J., Vendrell, M., Rossi, A. G., Kunath, T., Cockroft, S. L., Saleeb, R. S., Horrocks, M. H.Extracellular vesicles (EVs) are small, membrane-bound particles released by cells into the extracellular environment. They play a pivotal role in cell communication and have recently gained prominence as biomarkers. However, their low abundance and high heterogeneity challenges their accurate characterization using conventional approaches. To enable the specific detection of individual EVs, we coupled EV-specific antibodies labeled with two different fluorophores with fast-flow microfluidics and single-molecule confocal microscopy. This allowed us to determine the concentration of EVs down to femtomolar levels […]
- by Ghose, D., Nolen, J., Guan, K., Elston, T., Lew, D. J.Cells excel at interpreting noisy chemical gradients during fertilization, development, and immune responses, but the mechanisms underlying this remarkable ability remain poorly understood. Classical models for gradient sensing assume that information is derived solely from ligand-bound receptors, but cells can also utilize information from unbound receptors. Here, we first derive the physical limit for instantaneous gradient detection by a distribution of receptors on a sphere. Surprisingly, we find that a ratiometric biochemical strategy, which compares spatial distributions of active (bound) […]
- by de Vries, H. W., Barth, A., Fragasso, A., Otto, T. A., van der Graaf, A., van der Sluis, E. O., van der Giessen, E., Veenhoff, L. M., Dekker, C., Onck, P. R.Nucleocytoplasmic transport occurs via nuclear pore complexes (NPCs), ~40-60 nm wide pores lined with intrinsically disordered proteins that are rich in Phe-Gly motifs (FG-Nups) that form a selective barrier. Molecules larger than ~50 kDa are increasingly blocked for transport unless they are bound to a nuclear transport receptor (NTR). How the amino acid sequence of FG-Nups contribute to this is not fully understood. Here, we present de novo designed artificial FG-Nups with a systematically varied FG-repeat spacing and charge-to-hydrophobicity ratio […]
- by Deridoux, A., Heydari, S., Gorb, S., Kanso, E., Flammang, P., Gabriele, S.Sea stars use hundreds of tube feet on their oral surface to crawl, climb, and navigate complex environments, all without the coordination of a central brain. While the morphology of tube feet and their role as muscular hydrostats are well described, the dynamics underlying their locomotion remain poorly understood. To investigate these dynamics, we employed an optical imaging method based on frustrated total internal reflection to visualize and quantify tube foot adhesion in real time across individuals of Asterias rubens […]
- by Goyal, V., Grosh, K.Mammalian outer hair cells (OHCs) enhance sound amplification and frequency tuning through stereociliary hair bundles (HBs), which convert mechanical motion into electrical signals via mechano-electric transducer (MET) channels. Experiments show that the HB displacement creeps, and the MET current evinces dual timescales of adaptation in response to mechanical stimulus. Understanding these mechanisms is crucial for elucidating normal auditory function and disorders, yet their origins remain unclear. To address this, we developed a mathematical model of the OHC HB that incorporates […]
- by Hartmann, B., Reichel, F., Moeckel, C., Guck, J.The viscoelastic nature of biological cells has emerged as an increasingly important research subject due to its relevance for cellular functions under physiological and pathological conditions. Advancements in microfluidics have made this technology a promising tool to study the viscoelasticity of cells. However, significant challenges remain, including the complex distribution of stresses acting on cells depending on the channel geometry, and the difficulty of keeping cells in the focal plane for imaging. Here, we report a new approach using hyperbolic […]
- by Mandal, S., Chandrasekaran, A. R., Maiti, P. K.Designing biostable DNA nanocarriers for precise therapeutic delivery remains a key challenge in DNA nanotechnology due to susceptibility to nuclease degradation. Multi-stranded DNA nanostructures, such as Paranemic crossover (PX) DNA, show enhanced biostability compared to native duplex DNA due to their unique topology. However, the molecular origin of their exceptional nuclease resistance is still unknown. Using atomistic MD simulation and enhanced sampling, we uncover the molecular origins of PX-DNAs superior resistance over JX-DNA and dsDNA. Our findings reveal that PX-DNAs […]
- by Veit, S., Dearden, G. I., Menon, K. M., Noor, F., Menon, A. K., Pomorski, T. G.Scramblases play important roles in physiology by translocating phospholipids bidirectionally across cell membranes. For example, scrambling facilitated by VDAC1 dimers is the primary mechanism by which endoplasmic reticulum-derived phospholipids cross the outer membrane to enter mitochondria. Precise quantification of lipid scrambling, while critical for mechanistic understanding, cannot be obtained from ensemble averaged measurements of the activity of reconstituted scramblases. Here, we describe a microscopy platform for high-throughput imaging of single vesicles reconstituted with fluorescently-labeled phospholipids and heterogeneously crosslinked VDAC1 dimers. […]
- by Mandal, S., Chhetri, K. B., Jang, Y. H., Lansac, Y., Maiti, P. K.Protamine, an arginine-rich protein, compacts DNA more tightly than histones in somatic cells, yet its sequence-specific binding remains unclear. Using all-atom MD simulations with an arginine-rich short cationic peptide that mimics the protamine characteristics, we discovered distinct sequence preferences: the peptide binds preferentially to GC-rich sequences in the major groove and AT-rich sequences in the minor groove. Our structural analysis reveals that GC-rich binding induces significant DNA bending, narrowing the major groove and enhancing peptide interactions. In contrast, AT-rich minor […]
- by Vangheel, J., Ramon, H., Smeets, B.In morphogenesis and disease, biological tissues may exhibit diverse mechanical properties due to their capacity to switch between fluid-like to solid-like states through the (un)jamming transition. Here, we introduce a novel foam model to investigate how active mechanical properties and cellular interactions govern this transition in active cell monolayers. This model explicitly represents 3D cell shapes and describes cell-cell interactions via discrete interacting surfaces. Simulations reveal that cell-cell adhesive tension promotes tissue fluidization in high adhesive tissues, where it mainly […]
- by Cacheux, J., Quenan, T., Alcaide, D., Ordonez-Miranda, J., Jalabert, L., Nakano, S., Nakanishi, M., Cordelier, P., Bancaud, A., Matsunaga, Y. T.The poroelastic properties of tissues regulate molecular transport and mechanical signaling, yet their evolution during aging remains poorly understood. In particular, senescent fibroblasts accumulate in aged tissues, contributing to extracellular matrix (ECM) remodeling, but their impact on tissue mechanics and permeability is unclear. In this study, we developed a microfluidic-based in vitro model to assess the poroelastic properties of collagen gels embedded with senescent fibroblasts over time. Our approach integrates periodic pressure actuation with real-time pressure monitoring in a sealed […]
- by Camp, T., Li, Z., Li, Y., Oh, T.-J., Zhang, K.Oligomerization of photoactivatable proteins is widely used in optogenetics to modulate protein activity and regulate biological processes. However, their oligomerization states remain challenging to quantify in living cells. We applied photon counting histogram (PCH) analysis to quantify the oligomerization of two commonly used photoactivatable proteins, Vaucheria frigida Aureochrome light-oxygen-voltage (VfAuLOV) and Arabidopsis Thaliana cryptochrome 2 (AtCRY2), under dark and light conditions, providing a direct measurement of oligomerization states in live cells. Under blue light stimulation, VfAuLOV primarily forms dimers, whereas […]
- by Meng, K., Nie, L., Berger, J., von Grafenstein, N. R., Einholz, C., Rizzato, R., Schleicher, E., Bucher, D. B.Optically addressable spin systems, such as nitrogen-vacancy centers in diamond, have been widely studied for quantum sensing applications. In this work, we demonstrate that flavin-based cryptochrome proteins, which generate radical pairs upon optical excitation, also exhibit optically detected magnetic resonance. We further show that this optical spin interface is tunable by the protein structure. These findings establish radical pairs in proteins as a novel platform for optically addressable spin systems and magnetic field sensors. Additionally, the ability to control spin […]
- by Sohn, E. J., Sojitra, K. A., Rodrigues, L., Xu, X., Frost, B., Mittal, J., Libich, D. S.Ewing sarcoma, the second most common pediatric bone and soft tissue cancer, is caused by aberrant fusion of the EWS low-complexity domain (EWSLCD) to the DNA-binding domain of the transcription factor FLI1. The resulting fusion, EWS::FLI1, directly interacts with and engages in a dynamic interplay with EWS that drives tumorigenesis and regulates the function of both proteins. While EWSLCD is known to promote self-association, the role of the RNA-binding domains (RBDs) of EWS, which include RGG repeat regions and a […]
- by Marin, O., Kirchweger, P., Dalaloyan, A., Barak, Y., Elbaum, M.Protein condensation is the basis for formation of membrane-less organelles in the cell. Most famously, weak, polyvalent interactions, often including RNA, may lead to a liquid-liquid phase separation. This effect greatly enhances local concentrations and is thought to promote interactions that would remain rare in dilute solution. Synthetic systems provide a means to clarify the underlying biophysical mechanisms at play, both in vitro and in the cell via exogenous expression. In this regard, ferritin is a useful substrate, as its […]
- by Sandberg, J. W., Brannigan, G.The Envelope protein (E protein) of SARS-CoVs 1 and 2 has been implicated in the viral budding process and maintaining the spherical shape of the virus, but direct evidence linking the protein to long-range membrane deformation is still lacking. Computational predictions from molecular simulation have offered conflicting results, some showing long-range E-induced membrane curvature and others showing only local deformations. In the present study, we determine the mechanism driving these deformations by modulating the degree of hydrophobic mismatch between protein […]
- by Hur, M., Bartol, T., Rangamani, P., Sejnowski, T., Mjolsness, E.AO_SCPLOWBSTRACTC_SCPLOWThere is a morphodynamic component to synaptic learning by which changes in dendritic (postsynaptic) spine head size are associated with the strengthening or weakening of the synaptic connection between two neurons, in response to the temporal correlation of local presynaptic and postsynaptic signals. These morphological factors are in turn sculpted by the dynamics of the actin cytoskeleton. In this paper, we use Dynamical Graph Grammars (DGGs) implemented within a computer algebra system to model how networks of actin filaments can […]
- by Shen, M., Dayhoff, G. W., Shen, J.Protein ionization states provide electrostatic forces to modulate protein structure, stability, solubility, and function. Until now, predicting ionization states and understanding protein electrostatics have relied on structural information. Here we demonstrate that primary sequence alone enables remarkably accurate pK a predictions through KaML-ESM, a model that leverages evolutionary representations from ultra-large protein language models ESMs and pretraining with a synthetic pK a dataset. The KaML-ESM model achieves RMSEs approaching the experimental precision limit of [~]0.5 pH units for Asp, Glu, […]
- by Rahman, S., Morgan, M. T., Hicks, C. W., Gwizdala, A., Wolberger, C.Monoubiquitinated histone H2A lysine 119 (H2AK119ub) is a signature modification associated with transcriptional silencing and heterochromatin formation. Ubiquitin-specific protease 21 (USP21), one of four major deubiquitinating enzymes (DUBs) that target H2AK119ub, plays critical roles in diverse cellular processes1-4. The molecular mechanisms by which USP21 specifically deubiquitinates H2AK119ub and is regulated is unknown. USP21 contains a C-terminal USP catalytic domain, preceded by an N-terminal intrinsically disordered region (IDR). We determined the cryo-EM structure of the USP21 catalytic domain bound to an […]
- by Abelit, A. A., Boytsova, N. A., Kornev, A. A., Yakovleva, L. E., Daniil, S. D.In this paper, we aim to present a new intravital dye, called ABDS, which can be prepared using a marker pen and is useful for eukaryotic cell research. ABDS binds to the cytoplasmic membrane of living cells and can be used for their visualization in biophysical experiments, including Z-stack tomography, cell proliferation monitoring, and adhesion studies. Important properties of ABDS are its availability, bright stable fluorescence, manufacturing simplicity, and safety for living cells in vitro. The paper includes a method […]
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