PaperPlayer biorxiv biophysics

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.02.365098v1?rss=1

Authors: Plata, C. A., Marni, S., Maritan, A., Bellini, T., Suweis, S.

Abstract:

DNA hybridization is at the heart of countless biological and biotechnological processes. Its theoretical modeling played a crucial role, since it has enabled extracting the relevant thermodynamic parameters from systematic measurements of DNA melting curves. However, in its current state, hybridization modelling requires introducing an extra entropic contribution in self-complementary sequences that lacks any biophysical meaning. In this article, we propose a framework based on statistical physics to describe DNA hybridization and melting in an arbitrary mixture of DNA strands. In particular, we are able to analytically derive closed expressions of the system partition functions for any number N of strings, and explicitly calculate them in two paradigmatic situations: (i) a system made of self-complementary sequences and (ii) a system comprising two mutually complementary sequences. We derive the melting curve in the thermodynamic limit (N [->] {infty}) of our description, which differs from the expression commonly used to evaluate the melting of self-complementary systems in that it does not require correcting terms. We provide a thorough study comprising limit cases and alternative approaches showing how our framework can give a comprehensive view of hybridization and melting phenomena.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.04.367136v1?rss=1

Authors: Roy, U., Kwon, Y., Marie, L., Symington, L., Sung, P., Lisby, M., Greene, E. C.

Abstract:

Homologous recombination (HR) is essential for the maintenance of genome integrity. Rad51 paralogs fulfill a conserved, but undefined role in HR, and their mutations are associated with increased cancer risk in humans. Here, we use single-molecule imaging to reveal that the Saccharomyces cerevisiae Rad51 paralog complex Rad55-Rad57 promotes the assembly of Rad51 recombinase filaments through transient interactions, providing evidence that it acts as a classical molecular chaperone. Srs2 is an ATP-dependent anti-recombinase that downregulates HR by actively dismantling Rad51 filaments. Contrary to the current model, we find that Rad55-Rad57 does not physically block the movement of Srs2. Instead, Rad55-Rad57 promotes rapid re-assembly of Rad51 filaments after their disruption by Srs2. Our findings support a model in which Rad51 is in flux between free and ssDNA-bound states, the rate of which is dynamically controlled though the opposing actions of Rad55-Rad57 and Srs2.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.04.368837v1?rss=1

Authors: Liu, Y., Sun, M., Elnatan, D., Larson, A. G., Agard, D. A.

Abstract:

Hsp90 is a ubiquitous molecular chaperone that mediates the folding and maturation of hundreds of ''client'' proteins. Although Hsp90s generally function as homodimers, recent discoveries suggested that the mitochondrion specific Hsp90 (TRAP1) also forms functionally relevant tetramers. The structural mechanism of tetramer formation remains elusive. Here we used a combination of solution, biochemical and cryo-electron microscopy (cryo-EM) approaches to confirm that, independent of nucleotide state, a subpopulation of TRAP1 exists as tetramers. Unexpectedly, cryo-EM reveals multiple tetramer conformations having TRAP1 dimers arranged in orthogonal, parallel, or antiparallel configurations. The cryo-EM structure of one of the orthogonal tetrameric states was determined at 3.5 [A] resolution. Each of the two TRAP1 dimers is in a symmetric AMP{middle dot}PNP-bound closed state with the tetramer being stabilized through three distinct dimer-dimer interaction sites. In unique ways, each of the three TRAP1 domains contributes to tetramer formation. In addition to tetramerization via direct dimer-dimer contacts, our structure suggests that additional stabilization could come from domain swapping between the dimers. These results expand our understanding of TRAP1 biology beyond the conventional view of a functional dimer and provide a platform to further explore the function and regulation of tetrameric TRAP1 in mitochondria.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.04.368506v1?rss=1

Authors: Ryu, J.-K., Rah, S.-H., Janissen, R., Kerssemakers, J. W. J., Dekker, C.

Abstract:

The condensin SMC protein complex organizes chromosomal structure by extruding loops of DNA. Its ATP-dependent motor mechanism remains unclear but likely involves steps associated with large conformational changes within the ~50 nm protein complex. Here, using magnetic tweezers, we resolve single steps in loop extrusion by individual yeast condensins. Step sizes range between 20-45 nm at forces of 1.0-0.2 pN, respectively, comparable to the complex size. The large steps show that condensin reels in DNA in very sizeable amounts, up to ~600 bp per extrusion step, consistent with the non-stretched DNA polymer at these low forces. Using ATP-binding-impaired and ATP-hydrolysis-deficient mutants, we find that ATP binding is the primary step-generating stage underlying DNA loop extrusion. We discuss the findings in terms of a scrunching model where a stepwise DNA loop extrusion is generated by an ATP-binding-induced engagement of the hinge and the globular domain of the SMC complex.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.03.366179v1?rss=1

Authors: St Clair, J. R., Westacott, M. J., Farnsworth, N. L., Kravets, V., Schleicher, W. E., Miranda, J. G., Heintz, A., Ludin, N. W. F., Benninger, R. K.

Abstract:

Type2 diabetes results from failure of the {beta}-cell to compensate for insulin resistance, such as in obesity. Insulin secretion is governed by a series of metabolic and electrical events which can fail during the progression of diabetes. {beta}-cells are electrically coupled via Cx36 gap junction channels, thereby coordinating the pulsatile dynamics of electrical activity, Ca2+ and insulin release across the islet, enhancing insulin action. Pulsatile insulin release is disrupted in human type2 diabetes, although whether this disruption results from diminished gap junction coupling is unclear. Factors such as pro-inflammatory cytokines and free fatty acids disrupt gap junction coupling under invitro conditions. Here we test whether gap junction coupling and coordinated Ca2+ dynamics are disrupted in type2 diabetes, and whether recovery of gap junction coupling can recover islet function. We examine islets from healthy donors and those with type2 diabetes, as well as islets from db/db mice and islets treated with a cocktail of proinflammatory cytokines (TNF-, IL-1{beta}, IFN-{gamma}) or free fatty acids (palmitate). We modulate gap junction coupling using Cx36 over-expression or pharmacological activation via modafinil. We also develop a peptide mimetic (S293) of the c-terminal regulatory tail of Cx36 designed to compete against its phosphorylation and downregulation. Cx36 gap junction permeability and coordinated Ca2+ dynamics were disrupted in islets from human donors with type2 diabetes, as well as in islets from db/db mice or treated with proinflammatory cytokines or palmitate. Cx36 over-expression, modafinil treatment and S293 peptide all enhanced Cx36 gap junction coupling and protected against declines in coordinated Ca2+ dynamics. Cx36 over-expression and S293 peptide also reduced apoptosis induced by proinflammatory cytokines. Critically S293 peptide rescued gap junction coupling and Ca2+ dynamics in islets from both db/db mice and a sub-set of T2D donors. Thus, recovering or enhancing Cx36 gap junction coupling can improve islet function in diabetes.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.03.364851v1?rss=1

Authors: Lizarrondo, J., Klebl, D. P., Niebling, S., Abella, M., Schroer, M. A., Mertens, H. D. T., Veith, K., Svergun, D. I., Skruzny, M., Sobott, F., Muench, S., Garcia-Alai, M. M.

Abstract:

During clathrin-mediated endocytosis, a complex and dynamic network of protein-membrane interactions cooperate to achieve membrane invagination. Throughout this process, middle coat adaptors, Sla2 and Ent1, must remain attached to the plasma membrane to transmit force from the actin cytoskeleton required for successful membrane invagination. Here, we present a cryoEM structure of a 16-mer complex of membrane binding domains from Sla2 and Ent1 that anchors to the plasma membrane. Detailed mutagenesis in vitro and in vivo of the tetramer interfaces delineate the key interactions for complex formation and deficient cell growth phenotypes demonstrate the biological relevance of these interactions. Finally, time-resolved experiments in solution suggest that adaptors have evolved to achieve a fast subsecond timescale assembly in the presence of PIP2. Together, these findings provide a molecular understanding of an essential piece for the molecular puzzle of clathrin-coated sites.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.03.365106v1?rss=1

Authors: Reyer, M. A., Chennakesavalu, S., Heideman, E. M., Ma, X., Bujnowska, M., Hong, L., Dinner, A. R., Vanderpool, C. K., Fei, J.

Abstract:

Small RNAs (sRNAs) are important regulators of gene expression in bacteria, particularly during stress responses. Many genetically and biochemically well characterized sRNAs regulate gene expression post-transcriptionally, by affecting translation and degradation of the target mRNA after they bind to their targets through base pairing. However, how regulation at each of these levels quantitatively contributes to the overall efficacy of sRNA-mediated regulation is not well understood. Here we present a general approach combining imaging and mathematical modeling to determine kinetic parameters at different levels of sRNA-mediated gene regulation. Unexpectedly, our data reveal that certain previously characterized sRNAs are able to regulate some targets co-transcriptionally, rather strictly post-transcriptionally, and suggest that sRNA-mediated regulation can occur early in the mRNA's lifetime, perhaps as soon as the sRNA binding site is transcribed. In addition, our data suggest several important kinetic steps that may determine the efficiency and differential regulation of multiple mRNA targets by an sRNA. Particularly, binding of sRNA to the target mRNA is likely the rate-limiting step and may dictate the regulation hierarchy observed within an sRNA regulon.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.02.365825v1?rss=1

Authors: Schneider, M. M., Gautam, S., Herling, T. W., Andrzejwska, E., Krainer, G., Miller, A. M., Peter, Q. A. E., Ruggeri, F. S., Vendruscolo, M., Bracher, A., Dobson, C. M., Hartl, F.-U., Knowles, T.

Abstract:

Molecular chaperones contribute to the maintenance of cellular protein homeostasis through a wide range of mechanisms, including the assistance of de novo protein folding, the rescue of misfolded proteins, and the prevention of amyloid formation. Chaperones of the Hsp70 family have a striking capability of disaggregating otherwise irreversible aggregate structures such as amyloid fibrils that accumulate during the development of neurodegenerative diseases. However, the mechanisms of this key emerging functionality remain largely unknown. Here, we bring together microfluidic measurements with kinetic analysis and show that that the Hsp70 protein heat chock complement Hsc70 together with its two co-chaperones DnaJB1 and the nucleotide exchange factor Apg2 is able to completely reverse the aggregation process of alpha-synuclein, associated with Parkinson's disease, back to its soluble monomeric state. Moreover, we show that this reaction proceeds with first order kinetics in a process where monomer units are taken off directly from the fibril ends. Our results demonstrate that all components of the chaperone triad are essential for fibril disaggregation. Lastly, we quantify the interactions between the three chaperones as well as between the chaperones and the fibrils in solution, yielding both binding stoichiometries and dissociation constants. Crucially, we find that the stoichiometry of Hsc70 binding to fibrils suggests Hsc70 clustering at the fibril ends. Taken together, our results show that the mechanism of action of the Hsc70-DnaJB1-Apg2 chaperone system in disaggregating -synuclein fibrils involves the removal of monomer units without any intermediate fragmentation steps. These findings are fundamental to our understanding of the suppression of amyloid proliferation early in life and the natural clearance mechanisms of fibrillar deposits in Parkinson's disease, and inform on the possibilities and limitations of this strategy in the development of therapeutics against synucleinopathies and related neurodegenerative diseases.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.02.364513v1?rss=1

Authors: Maniou, E., Staddon, M., Marshall, A., Greene, N., Copp, A., Banerjee, S., Galea, G.

Abstract:

Gap closure is a common morphogenetic process. In mammals, failure to close the embryonic hindbrain neuropore (HNP) gap causes fatal anencephaly. We observed that surface ectoderm cells surrounding the mouse HNP assemble high-tension actomyosin purse-strings at their leading edge and establish the initial contacts across the embryonic midline. The HNP gap closes asymmetrically, faster from its rostral than caudal extreme, while maintaining an elongated aspect ratio. Cell-based physical modelling identifies two closure mechanisms sufficient to describe tissue-level HNP closure dynamics; purse-string contraction and directional cell crawling. Combining both closure mechanisms hastens gap closure and produces a constant rate of gap shortening. Purse-string contraction reduces, whereas crawling increases gap aspect ratio, and their combination maintains it. Closure rate asymmetry can be explained by embryo tissue geometry, namely a narrower rostral gap apex. At the cellular level, our model predicts highly directional cell migration with a constant rate of cells leaving the HNP rim. These behaviours are reproducibly live-imaged in mouse embryos. Thus, mammalian embryos coordinate cellular and tissue-level mechanics to achieve this critical gap closure event.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.11.01.364125v1?rss=1

Authors: Narla, A., Borenstein, D. B., Wingreen, N. S.

Abstract:

Bacteria grow on surfaces in complex immobile communities known as biofilms, which are composed of cells embedded in an extracellular matrix. Within biofilms, bacteria often interact with members of their own species, and cooperate or compete with members of other species via quorum sensing (QS). QS is a process by which microbes produce, secrete, and subsequently detect small molecules called autoinducers (AIs) to assess their local population density. We explore the competitive advantage of QS through agent-based simulations of a spatial model in which colony expansion via extracellular matrix production provides greater access to a limiting diffusible nutrient. We note a significant difference in results based on whether AI production is constitutive or limited by nutrient availability: If AI production is constitutive, simple QS-based matrix-production strategies can be far superior to any fixed strategy. However, if AI production is limited by nutrient availability, QS-based strategies fail to provide a significant advantage over fixed strategies. To explain this dichotomy, we derive a novel biophysical limit for the dynamic range of nutrient-limited AI concentrations in biofilms. This range is remarkably small (less than 10-fold) for the realistic case in which a growth-limiting diffusible nutrient is taken up within a narrow active growth layer. This biophysical limit implies that for QS to be most effective in biofilms, AI production should be a protected function not directly tied to metabolism.

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