PaperPlayer biorxiv biophysics

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.09.01.276568v1?rss=1

Authors: Owen, L. M., Bax, N. A., Weis, W., Dunn, A. R.

Abstract:

Focal adhesions (FAs) are large, integrin-based adhesion complexes that link cells to the extracellular matrix (ECM). Previous work demonstrates that FAs form only when and where they are necessary to transmit force between the cellular cytoskeleton and the ECM, but how this occurs remains poorly understood. Talin is a 270 kDa adapter protein that links integrins to filamentous (F)-actin and recruits additional components during FA assembly in a force-dependent manner. Cell biological and developmental data demonstrate that the third, and C-terminal, F-actin binding site (ABS3) of talin is required for normal FA formation. However, ABS3 binds F-actin only weakly in in vitro, biochemical assays. We used a single-molecule optical trap assay to examine how and whether ABS3 binds F-actin under physiologically relevant, pN mechanical loads. We find that ABS3 forms a directional catch bond with F-actin when force is applied towards the pointed end of the actin filament, with binding lifetimes more than 100-fold longer than when force is applied towards the barbed end. Long-lived bonds to F-actin under load require the ABS3 C-terminal dimerization domain, whose cleavage is known to regulate focal adhesion turnover. Our results support a mechanism in which talin ABS3 preferentially binds and orients actin filaments with barbed ends facing the cell periphery, thus nucleating long-range order in the actin cytoskeleton. We suggest that talin ABS3 may function as a molecular AND gate that allows FA growth only when sufficient integrin density, F-actin polarization, and mechanical tension are simultaneously present.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.31.276428v1?rss=1

Authors: Bauer, D., Ishikawa, H., Wemmer, K., Marshall, W. F.

Abstract:

Stochastic variations (noise) in gene expression have been extensively characterized, but the ramifications of this gene-level variation for cellular structure and function remain unclear. To what extent are cellular structures subject to noise? We show that flagellar length in Chlamydomonas exhibits significant variation that results from a combination of intrinsic fluctuations within the flagella and extrinsic cell to cell variation. We analyzed a series of candidate genes affecting flagella and found that flagellar length variation is increased in mutations which increase the average flagellar length, an effect that can be explained using a theoretical model for flagellar length regulation. Cells with greater differences in their flagellar lengths show impaired swimming but improved gliding motility, raising the possibility that cells have evolved mechanisms to tune intrinsic noise in length. Taken together our results show that biological noise exists at the level of subcellular structures, with a corresponding effect on cell function.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.09.01.277897v1?rss=1

Authors: Marx, D. C., Fleming, K. G.

Abstract:

Through the insertion of nonpolar side chains into the bilayer, the hydrophobic effect has long been accepted as a driving force for membrane protein folding. However, how the changing chemical composition of the bilayer affects the magnitude side chain transfer free energies ({Delta}G{degrees}sc) has historically not been well understood. A particularly challenging region for experimental interrogation is the bilayer interfacial region that is characterized by a steep polarity gradient. In this study we have determined the {Delta}G{degrees}sc for nonpolar side chains as a function of bilayer position using a combination of experiment and simulation. We discovered an empirical correlation between the surface area of nonpolar side chain, {Delta}G{degrees}sc, and local water concentration in the membrane that allows for {Delta}G{degrees}sc to be accurately estimated at any location in the bilayer. Using these water-to-bilayer {Delta}G{degrees}sc we have calculated the interface-to-bilayer transfer free energy ({Delta}G{degrees}(i,b)). We find that the {Delta}G{degrees}(i,b) are similar to the biological, translocon-based transfer free energies, indicating that the translocon energetically mimics the bilayer interface. Together these findings should increase the accuracy of computational workflows used to identify and design membrane proteins, as well as increase our understanding of how disease-causing mutations affect membrane protein folding and function.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.09.01.277186v1?rss=1

Authors: Gai, Y., Cook, B., Satru, S., Stone, H. A., Petry, S.

Abstract:

The organization of microtubules (MTs) is critical for cells during interphase and mitosis. During mitotic spindle assembly, MTs are made and organized around chromosomes in a process regulated by RanGTP. The role of RanGTP has been explored in Xenopus egg extracts, which are not limited by a cell membrane. Here, we investigated whether cell-sized confinements affect the assembly of RanGTP-induced MT networks in Xenopus egg extracts. We used microfluidics to encapsulate extracts within monodisperse extract-in-oil droplets. Importantly, we find that the architecture of Ran-induced MT networks depends on the droplet diameter and the Ran concentration, and differs from structures formed in bulk extracts. Our results highlight that both MT nucleation and physical confinement play critical roles in determining the spatial organization of the MT cytoskeleton.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.31.273466v1?rss=1

Authors: Yang, J., Mandriota, N., Harrellson, S. G., Jones-Molina, J. A., Yuste, R., Lefort, R., Sahin, O.

Abstract:

Synapses play a critical role in neural circuits, and they are potential sites for learning and memory. Maintenance of synaptic adhesion is critical for neural circuit function, however, biophysical mechanisms that help maintain synaptic adhesion are not clear. Studies with various cell types demonstrated the important role of stiffness in cellular adhesions. Although synaptic stiffness could also play a role in synaptic adhesion, stiffnesses of synapses are difficult to characterize due to their small size and challenges in verifying synapse identity and function. To address these challenges, we have developed an experimental platform that combines atomic force microscopy, fluorescence microscopy, and transmission electron microscopy. Here, using this platform, we report that functional, mature, excitatory synapses had an average elastic modulus of approximately 200 kPa, two orders of magnitude larger than that of the brain tissue, suggesting stiffness might have a role in synapse function. Similar to various functional and anatomical features of neural circuits, synaptic stiffness had a lognormal-like distribution, hinting a possible regulation of stiffness by processes involved in neural circuit function. In further support of this possibility, we observed that synaptic stiffness was correlated with spine size, a quantity known to correlate with synaptic strength. Using established stages of the long-term potentiation timeline and theoretical models of adhesion cluster dynamics, we developed a biophysical model of the synapse that not only explains extreme stiffness of synapses, their statistical distribution, and correlation with spine size, but also offers an explanation to how early biomolecular and structural changes during functional potentiation could lead to strengthening of synaptic adhesion. According to this model, synaptic stiffness serves as an indispensable physical messenger, feeding information back to synaptic adhesion molecules to facilitate maintenance of synaptic adhesion.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.29.273334v1?rss=1

Authors: Yousafzai, M. S., Yadav, V., Amiri, S., Staddon, M., Tabatabai, A. P., Errami, Y., Jaspard, G., Amiri, S., Banerjee, S., Murrell, M. P.

Abstract:

In liquid droplets, the balance of interfacial energies and substrate elasticity determines the shape of the droplet and the dynamics of wetting. In living cells, interfacial energies are not constant, but adapt to the mechanics of their environment. As a result, the forces driving the dynamics of wetting for cells and tissues are unclear and may be context specific. In this work, using a combination of experimental measurements and modeling, we show the surface tension of cell aggregates, as models of active liquid droplets, depends upon the size of the aggregate and the magnitude of applied load, which alters the wetting dynamics. Upon wetting rigid substrates, traction stresses are elevated at the boundary, and tension drives forward motion. By contrast, upon wetting compliant substrates, traction forces are attenuated, yet wetting occurs at a comparable rate. In this case, capillary forces at the contact line are elevated and aggregate surface tension contributes to strong outward, pressure-driven cellular flows. Thus, cell aggregates adapt to the mechanics of their environments, using pressure and traction as compensatory mechanisms to drive robust wetting.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.29.273755v1?rss=1

Authors: Zhang, C.-Y., Zhang, N.-H.

Abstract:

Structure and stiffness of nucleic acids are closely related to viral infectivity. The article is aimed to clarify a recent controversy on structure variation of viral DNA in temperature-dependent experiments. A multi-scale correlation between microscopic structure of viral DNA and its bulk stiffness is formulated by presenting a two-zone mesoscopic free energy model. Results reveal that the increasing temperature promotes DNA structure transformation from order to disorder, and leads to size effect on temperature-dependency of stiffness and structure of viral DNA.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.31.274860v1?rss=1

Authors: Lira, R. B., Leomil, F. S. C., Melo, R. J., Riske, K. d. A., Dimova, R.

Abstract:

Resealing of membrane pores is crucial for cell survival. We studied the membrane surface charge and medium composition as defining regulators triggering bursting and collapse of giant unilamellar vesicles upon poration. The pores were generated by either electric field or a detergent. Vesicles composed of mixtures of zwitterionic and negatively charged lipids at varying molar ratios were subjected to a single strong electric pulse and their response was characterized for a given membrane and medium composition. We observe that charged vesicles are prone to catastrophic vesicle collapse transforming them into tubular structures. The spectrum of destabilization responses includes the generation of long-living submicroscopic pores and partial vesicle bursting. The origin of these phenomena is related to the membrane edge tension, which governs pore closure. This edge tension significantly decreases as a function of the molar fraction of charged lipids. Destabilization of charged vesicles upon pore formation is a universal process since it is also observed with other poration stimuli. Disruption propensity is enhanced for membranes made of lipids with higher degree of unsaturation. It can be reversed by screening membrane charge in the presence of calcium ions. We interpret the observed findings in light of theories of stability and curvature generation in charged membranes and discuss mechanisms acting in cells to prevent total membrane collapse upon poration. Enhanced membrane stability is crucial for the success of electroporation-based technologies for cancer treatment and gene transfer.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.30.274340v1?rss=1

Authors: Nixon, C. F., Lim, S. A., Sailer, Z. R., Zheludev, I. N., Gee, C. L., Kelch, B. A., Harms, M. J., Marqusee, S.

Abstract:

In addition to encoding the final tertiary fold and stability, the primary sequence of a protein encodes the folding trajectory and kinetic barriers that determines the speed of folding. How these kinetic barriers are encoded by the sequence is not well understood. Here, we use evolutionary sequence variation in the alpha-lytic protease (aLP) protein family to probe the relationship between sequence and energy landscape. aLP has an unusual energy landscape: the native state of aLP is not the most thermodynamically favored conformation and, instead, it remains folded due to a large kinetic barrier preventing unfolding. In order to fold, aLP utilizes an N-terminal pro region of similar size to the protease itself that functions as a folding catalyst. Once folded, the pro region is removed, and the native state does not unfold on a biologically relevant timescale. Without the pro region, aLP folds on the order of millennia. A phylogenetic search uncovers aLP homologs with a wide range of pro-region sizes, including some with no pro region at all. In the resulting phylogenetic tree, these homologs cluster by pro-region size. Homologs naturally lacking pro regions are thermodynamically stable, fold much faster than aLP, yet retain the same fold as aLP. Key amino acids thought to contribute to aLP's extreme kinetic stability are lost in these homologs, further supporting their role in kinetic stability. This study highlights how the entire energy landscape plays an important role in determining the evolutionary pressures on and changes to the protein sequence.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.29.273474v1?rss=1

Authors: Liu, J., Hansen, D., Eck, E., Kim, Y. J., Turner, M. A., Alamos, S., Garcia, H. G.

Abstract:

The eukaryotic transcription cycle consists of three main steps: initiation, elongation, and cleavage of the nascent RNA transcript. Although each of these steps can be regulated as well as coupled with each other, their in vivo dissection has remained challenging because available experimental readouts lack sufficient spatiotemporal resolution to separate the contributions from each of these steps. Here, we describe a novel computational technique to simultaneously infer the effective parameters of the transcription cycle in real time and at the single-cell level using a two-color MS2/PP7 reporter gene and the developing fruit fly embryo as a case study. Our method enables detailed investigations into cell-to-cell variability in transcription-cycle parameters with high precision. These measurements, combined with theoretical modeling, reveal a significant variability in the elongation rate of individual RNA polymerase molecules. We further illustrate the power of this technique by uncovering a novel mechanistic connection between RNA polymerase density and nascent RNA cleavage efficiency. Thus, our approach makes it possible to shed light on the regulatory mechanisms in play during each step of the transcription cycle in individual, living cells at high spatiotemporal resolution.

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