Biology subjects

Wingreen, N. S.

Publications and source records attributed to Wingreen, N. S..

At least 55 records · Page 3Linked to original sources

bioRxiv2022

A rich get richer effect governs intracellular condensate size distributions

Phase separation of biomolecules into condensates has emerged as a ubiquitous mechanism for intracellular organization and impacts many intracellular processes, including reaction pathways through clustering of enzymes and their intermediates. Precise and rapid spatiotemporal control of reactions by condensates requires tuning of their sizes. However, the physical processes that govern the distribution of condensate sizes remain unclear. Here, we utilize a combination of synthetic and native condensates to probe the underlying physical mechanisms determining condensate size. We find that both native nuclear speckles and FUS condensates formed with the synthetic Corelet system obey an exponential size distribution, which can be recapitulated in Monte Carlo simulations of fast nucleation followed by coalescence. By contrast, pathological aggregation of cytoplasmic Huntingtin polyQ protein exhibits a power-law size distribution, with an exponent of -1.41 {+/-} 0.02. These distinct behaviors reflect the relative importance of nucleation and coalescence kinetics: introducing continuous condensate nucleation into the Monte Carlo coarsening simulations gives rise to polyQ-like power-law behavior. We demonstrate that the emergence of power-law distributions under continuous nucleation reflects a "rich get richer" effect, whose extent may play a general role in the determination of condensate size distributions.

biophysics↗

bioRxiv2021

Membrane fission during bacterial spore development requires DNA-driven cellular inflation

Bacteria require membrane fission for cell division and endospore formation. FisB catalyzes membrane fission during sporulation, but the molecular basis is unclear as it cannot remodel membranes by itself. Sporulation initiates with an asymmetric division that generates a large mother cell and a smaller forespore that contains only 1/4 of its complete genome. As the mother cell membranes engulf the forespore, a DNA translocase pumps the rest of the chromosome into the small forespore compartment, inflating it due to increased turgor. When the engulfing membranes undergo fission, the forespore is released into the mother cell cytoplasm. Here we show that forespore inflation and FisB accumulation are both required for efficient membrane fission. We suggest that high membrane tension in the engulfment membrane caused by forespore inflation drives FisB-catalyzed membrane fission. Collectively our data indicate that DNA-translocation has a previously unappreciated second function in energizing FisB-mediated membrane fission under energy-limited conditions. HIGHLIGHTS- Membrane fission during endospore formation requires rapid forespore inflation by ATP-driven DNA translocation. - Forespore inflation is fast enough to increase the tension of the engulfment and forespore membranes to near lysis tensions. - FisB catalyzes membrane fission by impeding the flux of lipids that partially support forespore and engulfment membrane growth. - Membrane fission utilizes chemical energy transduced to mechanical energy during DNA-packing into the forespore.

biophysics↗

bioRxiv2021

Quantitative input-output dynamics of a c-di-GMP signal-transduction cascade in Vibrio cholerae

Bacterial biofilms are multicellular communities that collectively overcome environmental threats and clinical treatments. To regulate the biofilm lifecycle, bacteria commonly transduce sensory information via the second-messenger molecule cyclic diguanylate (c-di-GMP). Using experimental and modeling approaches, we quantitatively capture c-di-GMP signal transmission via the bifunctional polyamine receptor NspS-MbaA, from ligand binding to output, in the pathogen Vibrio cholerae. Upon binding of norspermidine or spermidine, NspS-MbaA synthesizes or degrades c-di-GMP, respectively, which in turn, drives alterations specifically to biofilm gene expression. A longstanding question is how output specificity is achieved via c-di-GMP, a diffusible molecule that regulates dozens of effectors. We show that NspS-MbaA signals locally to specific effectors, sensitizing V. cholerae to polyamines. However, local signaling is not required for specificity, as changes to global cytoplasmic c-di-GMP levels can selectively regulate biofilm genes. This work establishes the input-output dynamics underlying c-di-GMP signaling, which could be useful for developing bacterial manipulation strategies.

microbiology↗

bioRxiv2021

piRNAs of Caenorhabditis elegans broadly silence nonself sequences through functionally random targeting

Small non-coding RNAs such as piRNAs serve as guides for Argonaute proteins, enabling sequence-specific, post-transcriptional regulation of gene expression. The piRNAs of Caenorhabditis elegans have been observed to bind targets with high mismatch tolerance, and appear to lack specific transposon targets, unlike piRNAs in Drosophila melanogaster and other organisms. These observations support a model in which C. elegans piRNAs provide a broad, indiscriminate net of silencing, acting in competition with siRNAs associated with the CSR-1 Argonaute that specifically protect self genes from silencing. However, the breadth of piRNA targeting has not been subject to in-depth quantitative analysis, nor has it been explained how piRNAs are distributed across sequence space to achieve complete coverage. Through a bioinformatic analysis of piRNA sequences, incorporating an original metric of piRNA-target distance, we demonstrate that C. elegans piRNAs are functionally random, in that their coverage of sequence space is comparable to that of random sequences. By possessing a sufficient number of distinct, essentially random piRNAs, C. elegans is able to target arbitrary nonself sequences with high probability. This result elucidates the mechanism by which newly transcribed mRNAs in C. elegans are classified as self or nonself, and has implications for piRNA evolution and biogenesis.

bioinformatics↗

bioRxiv2021

Noisy metabolism can drive the evolution of microbial cross-feeding

Cross-feeding, the exchange of nutrients between organisms, is ubiquitous in microbial communities. Despite its importance in natural and engineered microbial systems, our understanding of how cross-feeding arises is incomplete, with existing theories limited to specific scenarios. Here, we introduce a novel theory for the evolution of cross-feeding, which we term noise-averaging cooperation (NAC). NAC is based on the idea that, due to their small size, bacteria are prone to noisy regulation of metabolism which limits their growth rate. To compensate, related bacteria can share metabolites with each other to "average out" noise and improve their collective growth. This metabolite sharing among kin then allows for the evolution of metabolic interdependencies via gene deletions (this can be viewed as a generalization of the Black Queen Hypothesis). We first characterize NAC in a simple model of cell metabolism, showing that metabolite leakage can in principle substantially increase growth rate in a community context. Next, we develop a generalized framework for estimating the potential benefits of NAC among real bacteria. Using single-cell protein abundance data, we predict that bacteria suffer from substantial noise-driven growth inefficiencies, and may therefore benefit from NAC. Finally, we review existing evidence for NAC and outline potential experimental approaches to detect NAC in microbial communities.

microbiology↗

bioRxiv2021

Diffusion barriers and adaptive carbon uptake strategies enhance the modeled performance of the algal CO2-concentrating mechanism

Many photosynthetic organisms enhance the performance of their CO2-fixing enzyme Rubisco by operating a CO2-concentrating mechanism (CCM). Most CCMs in eukaryotic algae supply concentrated CO2 to Rubisco in an organelle called the pyrenoid. Ongoing efforts seek to engineer an algal CCM into crops that lack a CCM to increase yields. To advance our basic understanding of the algal CCM, we develop a chloroplast-scale reaction-diffusion model to analyze the efficacy and the energy efficiency of the CCM in the green alga Chlamydomonas reinhardtii. We show that achieving an effective and energetically efficient CCM requires a physical barrier such as thylakoid stacks or a starch sheath to reduce CO2 leakage out of the pyrenoid matrix. Our model provides insights into the relative performance of two distinct inorganic carbon uptake strategies: at air-level CO2, a CCM can operate effectively by taking up passively diffusing external CO2 and catalyzing its conversion to HCO3-, which is then trapped in the chloroplast; however, at lower external CO2 levels, effective CO2 concentration requires active import of HCO3-. We also find that proper localization of carbonic anhydrases can reduce futile carbon cycling between CO2 and HCO3-, thus enhancing CCM performance. We propose a four-step engineering path that increases predicted CO2 saturation of Rubisco up to seven-fold at a theoretical cost of only 1.5 ATP per CO2 fixed. Our system-level analysis establishes biophysical principles underlying the CCM that are broadly applicable to other algae and provides a framework to guide efforts to engineer an algal CCM into land plants. Significance StatementEukaryotic algae mediate approximately one-third of CO2 fixation in the global carbon cycle. Many algae enhance their CO2-fixing ability by operating a CO2-concentrating mechanism (CCM). Our model of the algal CCM lays a solid biophysical groundwork for understanding its operation. The models consistency with experimental observations supports existing hypotheses about the operating principles of the algal CCM and the functions of its component proteins. We provide a quantitative estimate of the CCMs energy efficiency and compare the performance of two distinct CO2 assimilation strategies under varied conditions. The model offers a quantitative framework to guide the engineering of an algal CCM into land plants and supports the feasibility of this endeavor.

biophysics↗

bioRxiv2020

Mechanical frustration of phase separation in the cell nucleus by chromatin

Liquid-liquid phase separation is a fundamental mechanism underlying subcellular organization. Motivated by the striking observation that optogenetically-generated droplets in the nucleus display suppressed coarsening dynamics, we study the impact of chromatin mechanics on droplet phase separation. We combine theory and simulation to show that crosslinked chromatin can mechanically suppress droplets coalescence and ripening, as well as quantitatively control their number, size, and placement. Our results highlight the role of the subcellular mechanical environment on condensate regulation.

biophysics↗

bioRxiv2020

A biophysical limit for quorum sensing in biofilms

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. Significance StatementBiofilms are a ubiquitous form of bacterial community. Within biofilms, bacteria communicate via chemical signals in a process called quorum sensing (QS). However, if signal production is nutrient-limited, then the nutrient-deficient interior of a biofilm cannot contribute to QS, which limits the ability of bacteria to assess their own population and behave accordingly. Numerical simulations of competitions among biofilm bacteria led us to discover a novel biophysical limit on the efficacy of nutrient-limited QS. In view of this limit, we conclude that to be most effective, QS signal production should be a prized function that is not metabolically slaved.

biophysics↗

bioRxiv2020

Sequence dependence of biomolecular phase separation

Cells are home to a wide variety of biomolecular condensates - phase-separated droplets that lack a membrane. In addition to nonspecific interactions, phase separation depends on specific binding motifs between constituent molecules. Nevertheless, few rules have been established on how these specific, heterotypic interactions drive phase separation. Using lattice-polymer simulations and mean-field theory, we show that the sequence of binding motifs strongly affects a polymers ability to phase separate, influencing both phase boundaries and condensate properties (e.g. viscosity and polymer diffusion). We find that sequences with large blocks of a single motif typically form more inter-polymer bonds which promote phase separation. Notably, the sequence of binding motifs influences phase separation primarily by determining the conformational entropy of self-bonding by single polymers. This contrasts with systems where the molecular architecture primarily affects the energy of the dense phase, providing a new entropy-based mechanism for the biological control of phase separation.

biophysics↗

bioRxiv2020

Decoding the physical principles of two-component biomolecular phase separation

Cells possess a multiplicity of non-membrane bound compartments, which form via liquid-liquid phase separation. These condensates assemble and dissolve as needed to enable central cellular functions. One important class of condensates is those composed of two associating polymer species that form one-to-one specific bonds. What are the physical principles that underlie phase separation in such systems? To address this question, we employed coarse-grained molecular dynamics simulations to examine how the phase boundaries depend on polymer valence, stoichiometry, and binding strength. We discovered a striking phenomenon - for sufficiently strong binding, phase separation is suppressed at rational polymer stoichiometries, which we termed the magic-ratio effect. We further developed an analytical dimer-gel theory that confirmed the magic-ratio effect and disentangled the individual roles of polymer properties in shaping the phase diagram. Our work provides new insights into the factors controlling the phase diagrams of biomolecular condensates, with implications for natural and synthetic systems.

biophysics↗

bioRxiv2020

The inner membrane protein YhdP modulates the rate of anterograde phospholipid flow in Escherichia coli

The outer membrane (OM) of Gram-negative bacteria is a selective permeability barrier that allows uptake of nutrients while simultaneously protecting the cell from harmful compounds. The basic pathways and molecular machinery responsible for transporting lipopolysaccharides (LPS), lipoproteins, and {beta}-barrel proteins to the OM have been identified, but very little is known about phospholipid (PL) transport. To identify genes capable of affecting PL transport, we screened for genetic interactions with mlaA*, a mutant in which anterograde PL transport causes the inner membrane (IM) to shrink and eventually rupture; characterization of mlaA*-mediated lysis suggested that PL transport can occur via a high-flux, diffusive flow mechanism. We found that YhdP, an IM protein involved in maintaining the OM permeability barrier, modulates the rate of PL transport during mlaA*-mediated lysis. Deletion of yhdP from mlaA* reduced the rate of IM transport to the OM by 50%, slowing shrinkage of the IM and delaying lysis. As a result, the weakened OM of {Delta}ydhP cells was further compromised and ruptured before the IM during mlaA*-mediated death. These findings demonstrate the existence of a high-flux, diffusive pathway for PL flow in Escherichia coli that is modulated by YhdP. Significance StatementThe outer membrane (OM) of Gram-negative bacteria serves as a barrier that protects cells from harmful chemical compounds, including many antibiotics. Understanding how bacteria build this barrier is an important step in engineering strategies to circumvent it. A long-standing mystery in the field is how phospholipids (PLs) are transported from the inner membrane (IM) to the OM. We previously discovered that a mutation in the gene mlaA causes rapid flow of PLs to the OM, eventually resulting in IM rupture. Here, we found that deletion of the gene yhdP delayed cell death in the mlaA mutant by slowing flow of PLs to the OM. These findings reveal a high-flux, diffusive pathway for PL transport in Gram-negative bacteria modulated by YhdP.

molecular biology↗

bioRxiv2020

Chromatin Mechanics Dictates Subdiffusion and Coarsening Dynamics of Embedded Condensates

DNA is organized into chromatin, a complex polymeric material which stores information and controls gene expression. An emerging mechanism for biological organization, particularly within the crowded nucleus, is biomolecular phase separation into condensed droplets of protein and nucleic acids. However, the way in which chromatin impacts the dynamics of phase separation and condensate formation is poorly understood. Here, we utilize a powerful optogenetic strategy to examine the interplay of droplet coarsening with the surrounding viscoelastic chromatin network. We demonstrate that droplet growth dynamics are directly inhibited by the chromatin-dense environment, which gives rise to an anomalously slow coarsening exponent, {beta}[~]0.12, contrasting with the classical prediction of {beta}[~] 1/3. Using scaling arguments and simulations, we show how this arrested growth can arise due to subdiffusion of individual condensates, predicting {beta}[~]/3, where is the diffusion exponent. Tracking the fluctuating motion of condensates within chromatin reveals a subdiffusive exponent, [~]0.5, which explains the anomalous coarsening behavior and is also consistent with Rouse-like dynamics arising from the entangled chromatin. Our findings have implications for the biophysical regulation of the size and shape of biomolecular condensates, and suggest that condensate emulsions can be used to probe the viscoelastic mechanical environment within living cells.

biophysics↗

bioRxiv2020

The Bacterial Cytoskeleton Spatially Confines Functional Membrane Microdomains

Cell membranes laterally segregate into microdomains enriched in certain lipids and scaffold proteins. Membrane microdomains modulate protein-protein interactions and are essential for cell polarity, signaling and membrane trafficking. How cells organize their membrane microdomains, and the physiological importance of these microdomains, is unknown. In eukaryotes, the cortical actin cytoskeleton is proposed to act like a fence, constraining the dynamics of membrane microdomains. Like their eukaryotic counterparts, bacterial cells have functional membrane microdomains (FMMs) that act as platforms for the efficient oligomerization of protein complexes. In this work, we used the model organism Bacillus subtilis to demonstrate that FMM organization and movement depend primarily on the interaction of FMM scaffold proteins with the domains protein cargo, rather than with domain lipids. Additionally, the MreB actin-like cytoskeletal network that underlies the bacterial membrane was found to frame areas of the membrane in which FMM mobility is concentrated. Variations in membrane fluidity did not affect FMM mobility whereas alterations in cell wall organization affected FMM mobility substantially. Interference with MreB organization alleviates FMM spatial confinement whereas, by contrast, inhibition of cell wall synthesis strengthens FMM confinement. The restriction of FMM lateral mobility by the submembranous actin-like cytoskeleton or the extracellular wall cytoskeleton appears to be a conserved mechanism in prokaryotic and eukaryotic cells for localizing functional protein complexes in specific membrane regions, thus contributing to the organization of cellular processes.

microbiology↗

bioRxiv2020

Competitive binding of independent extension and retraction motors explains the quantitative dynamics of type IV pili

The functions of type IV pili (TFP) are mediated by cycles of extension and retraction. The coordination of these cycles remains mysterious due to poor quantification of TFP dynamics. Here we fluorescently label the TFP in the opportunistic pathogen Pseudomonas aeruginosa and track the full extension and retraction cycles of individual TFP to quantify their dynamics. We test several models for the switch between extension and retraction using quantitative experiments, biophysical modeling and genetics. We invalidate the prominent hypothesis that this switch is triggered by surface contact. Instead, we show that the entire repetitive cycle of extension and retraction of individual TFP is governed by the stochastic binding of antagonistic extension and retraction motors and explain how this mechanism quantitatively defines physiologically-important features like TFP length and their production rate. Interestingly, our results suggest that the major throttle of TFP production is the unbinding of the retraction motor.

microbiology↗

bioRxiv2019

Spatial Ecology of Territorial Populations

Many ecosystems, from vegetation to biofilms, are composed of territorial populations that compete for both nutrients and physical space. What are the implications of such spatial organization for biodiversity? To address this question, we developed and analyzed a model of territorial resource competition. In the model, all species obey trade-offs inspired by biophysical constraints on metabolism; the species occupy non-overlapping territories while nutrients diffuse in space. We find that the nutrient diffusion time is an important control parameter for both biodiversity and the timescale of population dynamics. Interestingly, fast nutrient diffusion allows the populations of some species to fluctuate to zero, leading to extinctions. Moreover, territorial competition spontaneously gives rise to both multistability and the Allee effect (in which a minimum population is required for survival), so that small perturbations can have major ecological effects. While the assumption of trade-offs allows for the coexistence of more species than the number of nutrients - thus violating the principle of competitive exclusion - overall biodiversity is curbed by the domination of \"oligotroph\" species. Importantly, in contrast to well-mixed models, spatial structure renders diversity robust to inequalities in metabolic trade-offs. Our results suggest that territorial ecosystems can display high biodiversity and rich dynamics simply due to competition for resources in a spatial community.

ecology↗

bioRxiv2019

Modeling microbial diversity with metabolic trade-offs

Microbes face intense competition in the natural world, and so need to wisely allocate their resources to multiple functions, in particular to metabolism. Understanding competition among metabolic strategies that are subject to trade-offs is therefore crucial for deeper insight into the competition, cooperation, and community assembly of microorganisms. In this work, we evaluate competing metabolic strategies within an ecological context by considering not only how the environment influences cell growth, but also how microbes shape their chemical environment. Utilizing chemostat-based resource-competition models, we exhibit a set of intuitive and general procedures for assessing metabolic strategies. Using this framework, we are able to relate and unify multiple metabolic models, and to demonstrate how the fitness landscape of strategies becomes intrinsically dynamic due to species-environment feedback. Such dynamic fitness landscapes produce rich behaviors, and prove to be crucial for ecological and evolutionary stable coexistence in all the models we examined.

systems biology↗

bioRxiv2018

Revealing evolutionary constraints on proteins through sequence analysis

Alignments of large numbers of protein sequences have revealed \"sectors\" of collectively coevolving amino acids in several protein families. Here, we show that selection acting on any relevant physical property of a protein, e.g. the elastic energy of an important conformational change, can give rise to such a sector. We demonstrate that the main signature of these physical sectors lies in the smalleigenvalue modes of the covariance matrix of the selected sequences. This simple, generic model leads us to propose a principled method to identify sectors, along with the magnitudes of mutational effects, from sequence data.

biophysics↗

bioRxiv2018

Regulation of T cell expansion by antigen presentation dynamics

An essential feature of the adaptive immune system is the proliferation of antigen-specific lymphocytes during an immune reaction to form a large pool of effector cells. This proliferation must be regulated to ensure an effective response to infection while avoiding immunopathology. Recent experiments in mice have demonstrated that the expansion of a specific clone of T cells in response to cognate antigen obeys a striking inverse power law with respect to the initial number of T cells. Here, we show that such a relationship arises naturally from a model in which T cell expansion is limited by decaying levels of presented antigen. The same model also accounts for the observed dependence of T cell expansion on affinity for antigen and on the kinetics of antigen administration. Extending the model to address expansion of multiple T cell clones competing for antigen, we find that higher affinity clones can suppress the proliferation of lower affinity clones, thereby promoting the specificity of the response. Employing the model to derive optimal vaccination protocols, we find that exponentially increasing antigen doses can achieve a nearly optimized response. We thus conclude that the dynamics of presented antigen is a key regulator of both the size and specificity of the adaptive immune response.

immunology↗