Targeting the adaptability of heterogeneous aneuploids.

Aneuploid genomes, characterized by unbalanced chromosome stoichiometry (karyotype), are associated with cancer malignancy and drug resistance of pathogenic fungi. The phenotypic diversity resulting from karyotypic diversity endows the cell population with superior adaptability. We show here, using a combination of experimental data and a general stochastic model, that the degree of phenotypic variation, thus evolvability, escalates with the degree of overall growth suppression. Such scaling likely explains the challenge of treating aneuploidy diseases with a single stress-inducing agent. Instead, we propose the design of an "evolutionary trap" (ET) targeting both karyotypic diversity and fitness. This strategy entails a selective condition "channeling" a karyotypically divergent population into one with a predominant and predictably drugable karyotypic feature. We provide a proof-of-principle case in budding yeast and demonstrate the potential efficacy of this strategy toward aneuploidy-based azole resistance in Candida albicans. By analyzing existing pharmacogenomics data, we propose the potential design of an ET against glioblastoma.

Recombination between heterologous human acrocentric chromosomes.

The short arms of the human acrocentric chromosomes 13, 14, 15, 21 and 22 (SAACs) share large homologous regions, including ribosomal DNA repeats and extended segmental duplications1,2. Although the resolution of these regions in the first complete assembly of a human genome-the Telomere-to-Telomere Consortium's CHM13 assembly (T2T-CHM13)-provided a model of their homology3, it remained unclear whether these patterns were ancestral or maintained by ongoing recombination exchange. Here we show that acrocentric chromosomes contain pseudo-homologous regions (PHRs) indicative of recombination between non-homologous sequences. Utilizing an all-to-all comparison of the human pangenome from the Human Pangenome Reference Consortium4 (HPRC), we find that contigs from all of the SAACs form a community. A variation graph5 constructed from centromere-spanning acrocentric contigs indicates the presence of regions in which most contigs appear nearly identical between heterologous acrocentric chromosomes in T2T-CHM13. Except on chromosome 15, we observe faster decay of linkage disequilibrium in the pseudo-homologous regions than in the corresponding short and long arms, indicating higher rates of recombination6,7. The pseudo-homologous regions include sequences that have previously been shown to lie at the breakpoint of Robertsonian translocations8, and their arrangement is compatible with crossover in inverted duplications on chromosomes 13, 14 and 21. The ubiquity of signals of recombination between heterologous acrocentric chromosomes seen in the HPRC draft pangenome suggests that these shared sequences form the basis for recurrent Robertsonian translocations, providing sequence and population-based confirmation of hypotheses first developed from cytogenetic studies 50 years ago9.

Motility and segregation of Hsp104-associated protein aggregates in budding yeast.

During yeast cell division, aggregates of damaged proteins are segregated asymmetrically between the bud and the mother. It is thought that protein aggregates are cleared from the bud via actin cable-based retrograde transport toward the mother and that Bni1p formin regulates this transport. Here, we examined the dynamics of Hsp104-associated protein aggregates by video microscopy, particle tracking, and image correlation analysis. We show that protein aggregates undergo random walk without directional bias. Clearance of heat-induced aggregates from the bud does not depend on formin proteins but occurs mostly through dissolution via Hsp104p chaperon. Aggregates formed naturally in aged cells also exhibit random walk but do not dissolve during observation. Although our data do not disagree with a role for actin or cell polarity in aggregate segregation, modeling suggests that their asymmetric inheritance can be a predictable outcome of aggregates' slow diffusion and the geometry of yeast cells.

Control of the mitotic cleavage plane by local epithelial topology.

For nearly 150 years, it has been recognized that cell shape strongly influences the orientation of the mitotic cleavage plane (e.g., Hofmeister, 1863). However, we still understand little about the complex interplay between cell shape and cleavage-plane orientation in epithelia, where polygonal cell geometries emerge from multiple factors, including cell packing, cell growth, and cell division itself. Here, using mechanical simulations, we show that the polygonal shapes of individual cells can systematically bias the long-axis orientations of their adjacent mitotic neighbors. Strikingly, analyses of both animal epithelia and plant epidermis confirm a robust and nearly identical correlation between local cell topology and cleavage-plane orientation in vivo. Using simple mathematics, we show that this effect derives from fundamental packing constraints. Our results suggest that local epithelial topology is a key determinant of cleavage-plane orientation, and that cleavage-plane bias may be a widespread property of polygonal cell sheets in plants and animals.

Phase plane dynamics of ERK phosphorylation.

The extracellular signal-regulated kinase (ERK) controls multiple critical processes in the cell and is deregulated in human cancers, congenital abnormalities, immune diseases, and neurodevelopmental syndromes. Catalytic activity of ERK requires dual phosphorylation by an upstream kinase, in a mechanism that can be described by two sequential Michaelis-Menten steps. The estimation of individual reaction rate constants from kinetic data in the full mechanism has proved challenging. Here, we present an analytically tractable approach to parameter estimation that is based on the phase plane representation of ERK activation and yields two combinations of six reaction rate constants in the detailed mechanism. These combinations correspond to the ratio of the specificities of two consecutive phosphorylations and the probability that monophosphorylated substrate does not dissociate from the enzyme before the second phosphorylation. The presented approach offers a language for comparing the effects of mutations that disrupt ERK activation and function in vivo. As an illustration, we use phase plane representation to analyze dual phosphorylation under heterozygous conditions, when two enzyme variants compete for the same substrate.

Quartz Crystal Microbalance Frequency Response to Discrete Adsorbates in Liquids.

Quartz crystal microbalance with dissipation monitoring (QCM-D) has become a major tool enabling accurate investigation of the adsorption kinetics of nanometric objects such as DNA fragments, polypeptides, proteins, viruses, liposomes, polymer, and metal nanoparticles. However, in liquids, a quantitative analysis of the experimental results is often intricate because of the complex interplay of hydrodynamic and adhesion forces varying with the physicochemical properties of adsorbates and functionalized QCM-D sensors. In the present paper, we dissect the role of hydrodynamics for the analytically tractable case of stiff contact, whereas the adsorbed rigid particles oscillate with the resonator without rotation. Under the assumption of the low surface coverage, we theoretically study the excess shear force exerted on the resonator, which has two contributions: (i) the fluid-mediated force due to flow disturbance created by the particle and (ii) the force exerted on the particle by the fluid and transmitted to the sensor via contact. The theoretical analysis enables an accurate interpretation of the QCM-D impedance measurements. It is demonstrated inter alia that for particles of the size comparable with protein molecules, the hydrodynamic force dominates over the inertial force and that the apparent mass derived from QCM independently of the overtone is about 10 times the Sauerbrey (inertial) mass. The theoretical results show excellent agreement with the results of experiments and advanced numerical simulations for a wide range of particle sizes and oscillation frequencies.

Pushing myelination - developmental regulation of myosin expression drives oligodendrocyte morphological differentiation.

Oligodendrocytes are the central nervous system myelin-forming cells providing axonal electrical insulation and higher-order neuronal circuitry. The mechanical forces driving the differentiation of oligodendrocyte precursor cells into myelinating oligodendrocytes are largely unknown, but likely require the spatiotemporal regulation of the architecture and dynamics of the actin and actomyosin cytoskeletons. In this study, we analyzed the expression pattern of myosin motors during oligodendrocyte development. We report that oligodendrocyte differentiation is regulated by the synchronized expression and non-uniform distribution of several members of the myosin network, particularly non-muscle myosins 2B and 2C, which potentially operate as nanomechanical modulators of cell tension and myelin membrane expansion at different cell stages.This article has an associated First Person interview with the first author of the paper.

Calculating absorption dose when X-ray irradiation modifies material quantity and chemistry.

X-ray absorption is a sensitive and versatile tool for chemical speciation. However, when high doses are used, the absorbed energy can change the composition, amount and structure of the native material, thereby changing the aspects of the absorption process on which speciation is based. How can one calculate the dose when X-ray irradiation affects the chemistry and changes the amount of the material? This paper presents an assumption-free approach which can retrieve from the experimental data all dose-sensitive parameters - absorption coefficients, composition (elemental molecular units), material densities - which can then be used to calculate accurate doses as a function of irradiation. This approach is illustrated using X-ray damage to a solid film of a perfluorosulfonic acid fluoropolymer in a scanning transmission soft X-ray microscope. This new approach is compared against existing dose models which calculate the dose by making simplifying assumptions regarding the material quantity, density and chemistry. While the detailed measurements used in this approach go beyond typical methods to experimental analytical X-ray absorption, they provide a more accurate quantitation of radiation dose, and help to understand mechanisms of radiation damage.

Fluid-Mediated Force on a Particle Due to an Oscillating Plate and Its Effect on Deposition Measurements by a Quartz Crystal Microbalance.

Interaction of particles with boundaries is a fundamental problem in many fields of physics. In this Letter, we theoretically examine the fluid-mediated interaction between a horizontally oscillating plate and a spherical particle, revealing emergence of the novel nonlinear vertical force exerted on the particle. Although we demonstrate that the phenomenon only slightly alters deposition of colloidal (sub-)µm-sized particles measured by quartz crystal microbalance, it can result in levitation of larger particles above the plate, considerably hindering their deposition.

Protrusion membrane pearling emerges during 3D cell division.

Cell division is accompanied by dramatic changes in shape that ultimately lead to the physical separation of one cell into two. In 2D microenvironments, cells round up and remain adhered onto the substrate by thin retraction fibers during division. In contrast, in 3D environments, cells divide exhibiting long protrusions that guide the orientation of the division axis. However, the mechanism of cell division in three dimensions still remains poorly understood. Here we report the spontaneous formation of transient quasiperiodic membrane pearling on extended mitotic protrusions during 3D cell division. Protrusion membrane pearling may be initiated by the non-uniform distribution of focal adhesions and consequent stationary instability of the protrusive membrane. Overall, membrane pearling emergence may provide insights into a novel modality of 3D cell division with potential physiological relevance.

Mechanical plasticity during oligodendrocyte differentiation and myelination.

In the central nervous system, oligodendrocyte precursor cells are exclusive in their potential to differentiate into myelinating oligodendrocytes. Oligodendrocyte precursor cells migrate within the parenchyma and extend cell membrane protrusions that ultimately evolve into myelinating sheaths able to wrap neuronal axons and significantly increase their electrical conductivity. The subcellular force generating mechanisms driving morphological and functional transformations during oligodendrocyte differentiation and myelination remain elusive. In this review, we highlight the mechanical processes governing oligodendrocyte plasticity in a dynamic interaction with the extracellular matrix.

First-principles X-ray absorption dose calculation for time-dependent mass and optical density.

A dose integral of time-dependent X-ray absorption under conditions of variable photon energy and changing sample mass is derived from first principles starting with the Beer-Lambert (BL) absorption model. For a given photon energy the BL dose integral D(e, t) reduces to the product of an effective time integral T(t) and a dose rate R(e). Two approximations of the time-dependent optical density, i.e. exponential A(t) = c + aexp(-bt) for first-order kinetics and hyperbolic A(t) = c + a/(b + t) for second-order kinetics, were considered for BL dose evaluation. For both models three methods of evaluating the effective time integral are considered: analytical integration, approximation by a function, and calculation of the asymptotic behaviour at large times. Data for poly(methyl methacrylate) and perfluorosulfonic acid polymers measured by scanning transmission soft X-ray microscopy were used to test the BL dose calculation. It was found that a previous method to calculate time-dependent dose underestimates the dose in mass loss situations, depending on the applied exposure time. All these methods here show that the BL dose is proportional to the exposure time D(e, t) ≃ K(e)t.

Myosin Clusters of Finite Size Develop Contractile Stress in 1D Random Actin Arrays.

Myosin-powered force generation and contraction in nonmuscle cells underlies many cell biological processes and is based on contractility of random actin arrays. This contractility must rely on a microscopic asymmetry, the precise mechanism of which is not completely clear. A number of models of mechanical and structural asymmetries in actomyosin contraction have been posited. Here, we examine a contraction mechanism based on a finite size of myosin clusters and anisotropy of force generation by myosin heads at the ends of the myosin clusters. We use agent-based numerical simulations to demonstrate that if average lengths of actin filaments and myosin clusters are similar, then the proposed microscopic asymmetry leads to effective contraction of random 1D actomyosin arrays. We discuss the model's implication for mechanics of contractile rings and stress fibers.

A Combination of Actin Treadmilling and Cross-Linking Drives Contraction of Random Actomyosin Arrays.

We investigate computationally the self-organization and contraction of an initially random actomyosin ring. In the framework of a detailed physical model for a ring of cross-linked actin filaments and myosin-II clusters, we derive the force balance equations and solve them numerically. We find that to contract, actin filaments have to treadmill and to be sufficiently cross linked, and myosin has to be processive. The simulations reveal how contraction scales with mechanochemical parameters. For example, they show that the ring made of longer filaments generates greater force but contracts slower. The model predicts that the ring contracts with a constant rate proportional to the initial ring radius if either myosin is released from the ring during contraction and actin filaments shorten, or if myosin is retained in the ring, while the actin filament number decreases. We demonstrate that a balance of actin nucleation and compression-dependent disassembly can also sustain contraction. Finally, the model demonstrates that with time pattern formation takes place in the ring, worsening the contractile process. The more random the actin dynamics are, the higher the contractility will be.

On the origins of the mitotic shift in proliferating cell layers.

BACKGROUND: During plant and animal development, monolayer cell sheets display a stereotyped distribution of polygonal cell shapes. In interphase cells these shapes range from quadrilaterals to decagons, with a robust average of six sides per cell. In contrast, the subset of cells in mitosis exhibits a distinct distribution with an average of seven sides. It remains unclear whether this 'mitotic shift' reflects a causal relationship between increased polygonal sidedness and increased division likelihood, or alternatively, a passive effect of local proliferation on cell shape. METHODS: We use a combination of probabilistic analysis and mathematical modeling to predict the geometry of mitotic polygonal cells in a proliferating cell layer. To test these predictions experimentally, we use Flp-Out stochastic labeling in the Drosophila wing disc to induce single cell clones, and confocal imaging to quantify the polygonal topologies of these clones as a function of cellular age. For a more generic test in an idealized cell layer, we model epithelial sheet proliferation in a finite element framework, which yields a computationally robust, emergent prediction of the mitotic cell shape distribution. RESULTS: Using both mathematical and experimental approaches, we show that the mitotic shift derives primarily from passive, non-autonomous effects of mitoses in neighboring cells on each cell's geometry over the course of the cell cycle. Computationally, we predict that interphase cells should passively gain sides over time, such that cells at more advanced stages of the cell cycle will tend to have a larger number of neighbors than those at earlier stages. Validating this prediction, experimental analysis of randomly labeled epithelial cells in the Drosophila wing disc demonstrates that labeled cells exhibit an age-dependent increase in polygonal sidedness. Reinforcing these data, finite element simulations of epithelial sheet proliferation demonstrate in a generic framework that passive side-gaining is sufficient to generate a mitotic shift. CONCLUSIONS: Taken together, our results strongly suggest that the mitotic shift reflects a time-dependent accumulation of shared cellular interfaces over the course of the cell cycle. These results uncover fundamental constraints on the relationship between cell shape and cell division that should be general in adherent, polarized cell layers.

Whole chromosome aneuploidy: big mutations drive adaptation by phenotypic leap.

Despite its widespread existence, the adaptive role of aneuploidy (the abnormal state of having an unequal number of different chromosomes) has been a subject of debate. Cellular aneuploidy has been associated with enhanced resistance to stress, whereas on the organismal level it is detrimental to multicellular species. Certain aneuploid karyotypes are deleterious for specific environments, but karyotype diversity in a population potentiates adaptive evolution. To reconcile these paradoxical observations, this review distinguishes the role of aneuploidy in cellular versus organismal evolution. Further, it proposes a population genetics perspective to examine the behavior of aneuploidy on a populational versus individual level. By altering the copy number of a significant portion of the genome, aneuploidy introduces large phenotypic leaps that enable small cell populations to explore a wide phenotypic landscape, from which adaptive traits can be selected. The production of chromosome number variation can be further increased by stress- or mutation-induced chromosomal instability, fueling rapid cellular adaptation.

Modular coherence of protein dynamics in yeast cell polarity system.

In this study, we investigated on a systems level how complex protein interactions underlying cell polarity in yeast determine the dynamic association of proteins with the polar cortical domain (PCD) where they localize and perform morphogenetic functions. We constructed a network of physical interactions among >100 proteins localized to the PCD. This network was further divided into five robust modules correlating with distinct subprocesses associated with cell polarity. Based on this reconstructed network, we proposed a simple model that approximates a PCD protein's molecular residence time as the sum of the characteristic time constants of the functional modules with which it interacts, weighted by the number of edges forming these interactions. Regression analyses showed excellent fitting of the model with experimentally measured residence times for a large subset of the PCD proteins. The model is able to predict residence times using small training sets. Our analysis also revealed a scaffold protein that imposes a local constraint of dynamics for certain interacting proteins.

Modeling the evolution of Schizosaccharomyces pombe populations with multiple killer meiotic drivers.

Meiotic drivers are selfish genetic loci that can be transmitted to more than half of the viable gametes produced by a heterozygote. This biased transmission gives meiotic drivers an evolutionary advantage that can allow them to spread over generations until all members of a population carry the driver. This evolutionary power can also be exploited to modify natural populations using synthetic drivers known as "gene drives." Recently, it has become clear that natural drivers can spread within genomes to birth multicopy gene families. To understand intragenomic spread of drivers, we model the evolution of 2 or more distinct meiotic drivers in a population. We employ the wtf killer meiotic drivers from Schizosaccharomyces pombe, which are multicopy in all sequenced isolates, as models. We find that a duplicate wtf driver identical to the parent gene can spread in a population unless, or until, the original driver is fixed. When the duplicate driver diverges to be distinct from the parent gene, we find that both drivers spread to fixation under most conditions, but both drivers can be lost under some conditions. Finally, we show that stronger drivers make weaker drivers go extinct in most, but not all, polymorphic populations with absolutely linked drivers. These results reveal the strong potential for natural meiotic drive loci to duplicate and diverge within genomes. Our findings also highlight duplication potential as a factor to consider in the design of synthetic gene drives.

Positional information modulates transient regeneration-activated cell states during vertebrate appendage regeneration.

Injury is common in the life of organisms. Because the extent of damage cannot be predicted, injured organisms must determine how much tissue needs to be restored. Although it is known that amputation position affects the regeneration speed of appendages, mechanisms conveying positional information remain unclear. We investigated tissue dynamics in regenerating caudal fins of the African killifish (Nothobranchius furzeri) and found position-specific, differential spatial distribution modulation, persistence, and magnitude of proliferation. Single-cell RNA sequencing revealed a transient regeneration-activated cell state (TRACS) in the basal epidermis that is amplified to match a given amputation position and expresses components and modifiers of the extracellular matrix (ECM). Notably, CRISPR-Cas9-mediated deletion of the ECM modifier sequestosome 1 (sqstm1) increased the regenerative capacity of distal injuries, suggesting that regeneration growth rate can be uncoupled from amputation position. We propose that basal epidermis TRACS transduce positional information to the regenerating blastema by remodeling the ECM.

Force to divide: structural and mechanical requirements for actomyosin ring contraction.

One of the unresolved questions in the field of cell division is how the actomyosin cytoskeleton remains structurally organized while generating the contractile force to divide one cell into two. In analogy to the actomyosin-based force production mechanism in striated muscle, it was originally proposed that contractile stress in the actomyosin ring is generated via a sliding filament mechanism within an organized sarcomere-like array. However, over the last 30 years, ultrastructural and functional studies have noted important distinctions between cytokinetic structures in dividing cells and muscle sarcomeres. Myosin-II motor activity is not always required, and there is evidence that actin depolymerization contributes to contraction. In this Review, the architecture and contractile dynamics of the actomyosin ring at the cell division plane will be discussed. We will report the interdisciplinary advances in the field as well as their integration into a mechanistic understanding of contraction in cell division and in other biological processes that rely on an actomyosin-based force-generating system.