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Activation of myosin light-chain kinase (MLCK) by calcium ions (Ca2+) and calmodulin (CaM) plays an important role in numerous cellular functions including vascular smooth muscle contraction and cellular motility. Despite extensive biochemical analysis of this system, aspects of the mechanism of activation remain controversial, and competing theoretical models have been proposed for the binding of Ca2+and CaM to MLCK. The models are analytically solvable for an equilibrium steady state and give rise to distinct predictions that hold regardless of the numerical values assigned to parameters. These predictions form the basis of a recently proposed, multi-part experimental strategy for model discrimination. Here we implement this strategy by measuring CaM-MLCK binding using anin vitroFRET system. This system uses the CaM-binding region of smooth muscle MLCK protein to link two fluorophores to form an MLCK FRET Reporter (FR). Biochemical and biophysical experiments have established that FR can be reliably used to analyze MLCK-CaM binding. We assessed the binding of either wild-type CaM, or mutant CaM with one or more defective EF-hand domains, to FR. Interpretation of binding data in light of the mathematical models suggests a partially ordered mechanism for binding of CaM to MLCK. Complementary data collected using orthogonal approaches that directly quantify CaM-MLCK binding further supports our conclusions.

Lack of high-quality multilingual resources can contribute to disparities in the availability of medical and public health information. The COVID-19 pandemic has required rapid dissemination of essential guidance to diverse audiences and therefore provides an ideal context in which to study linguistic fairness in the U.S. Here we report a cross-sectional study of official non-English information about COVID-19 from the Centers for Disease Control and Prevention, the Food and Drug Administration, and the health departments of all 50 U.S. states. We find that multilingual information is limited in many states, such that almost half of all individuals not proficient in English or Spanish lack access to state-specific COVID-19 guidance in their primary language. Although Spanish-language information is widely available, we show using automated readability formulas that most materials do not follow standard recommendations for clear communication in medicine and public health. In combination, our results provide a snapshot of linguistic unfairness across the U.S. and highlight an urgent need for the creation of plain language, multilingual resources about COVID-19.

On April 13, 2021 the Centers for Disease Control and Prevention (CDC) and Food and Drug Administration (FDA) recommended in a pause in use of the Johnson & Johnson (J&J)-Janssen COVID-19 vaccine due to reports of cerebral venous sinus thrombosis (CVST) in recently vaccinated individuals. The announcement of the pause required development of a coordinated communication strategy under extreme time pressure and careful messaging by stakeholders to mitigate reduced public confidence in COVID-19 vaccines, as was observed following the temporary suspension of the Oxford-AstraZeneca vaccine in many countries. In this survey study, we evaluated understanding and impressions of the CDC’s public online information about the J&J-Janssen pause among unvaccinated US adults.

The corpus of Old English verse is an indispensable source for scholars of the Indo-European tradition, early Germanic culture and English literary history. Although it has been the focus of sustained literary scholarship for over two centuries, Old English poetry has not been subjected to corpus-wide computational profiling, in part because of the sparseness and extreme fragmentation of the surviving material. Here we report a detailed quantitative analysis of the whole corpus that considers a broad range of features reflective of sound, metre and diction. This integrated examination of fine-grained features enabled us to identify salient stylistic patterns, despite the inherent limitations of the corpus. In particular, we provide quantitative evidence consistent with the unitary authorship of Beowulf and the Cynewulfian authorship of Andreas, shedding light on two longstanding questions in Old English philology. Our results demonstrate the usefulness of high-dimensional stylometric profiling for fragmentary literary traditions and lay the foundation for future studies of the cultural evolution of English literature.

Identifying the stylistic signatures characteristic of different genres is of central importance to literary theory and criticism. In this article we report a large-scale computational analysis of Latin prose and verse using a combination of quantitative stylistics and supervised machine learning. We train a set of classifiers to differentiate prose and poetry with high accuracy (>97%) based on a set of twenty-six text-based, primarily syntactic features and rank the relative importance of these features to identify a low-dimensional set still sufficient to achieve excellent classifier performance. This analysis demonstrates that Latin prose and verse can be classified effectively using just three top features. From examination of the highly ranked features, we observe that measures of the hypotactic style favored in Latin prose (i.e. subordinating constructions in complex sentences, such as relative clauses) are especially useful for classification.

Identifying intertextual relationships between authors is of central importance to the study of literature. We report an empirical analysis of intertextuality in classical Latin literature using word embedding models. To enable quantitative evaluation of intertextual search methods, we curate a new dataset of 945 known parallels drawn from traditional scholarship on Latin epic poetry. We train an optimized word2vec model on a large corpus of lemmatized Latin, which achieves state-of-the-art performance for synonym detection and outperforms a widely used lexical method for intertextual search. We then demonstrate that training embeddings on very small corpora can capture salient aspects of literary style and apply this approach to replicate a previous intertextual study of the Roman historian Livy, which relied on hand-crafted stylometric features. Our results advance the development of core computational resources for a major premodern language and highlight a productive avenue for cross-disciplinary collaboration between the study of literature and NLP.

This cross-sectional study evaluates the readability of information on COVID-19 on websites of international agencies, governments of 15 countries, and US health departments.

Summary Complex behavior is associated with animals with nervous systems, but decision-making and learning also occur in non-neural organisms [ 1 ], including singly nucleated cells [ 2 , 3 , 4 , 5 ] and multi-nucleate synctia [ 6 , 7 , 8 ]. Ciliates are single-cell eukaryotes, widely dispersed in aquatic habitats [ 9 ], with an extensive behavioral repertoire [ 10 , 11 , 12 , 13 ]. In 1906, Herbert Spencer Jennings [ 14 , 15 ] described in the sessile ciliate Stentor roeseli a hierarchy of responses to repeated stimulation, which are among the most complex behaviors reported for a singly nucleated cell [ 16 , 17 ]. These results attracted widespread interest [ 18 , 19 ] and exert continuing fascination [ 7 , 20 , 21 , 22 ] but were discredited during the behaviorist orthodoxy by claims of non-reproducibility [ 23 ]. These claims were based on experiments with the motile ciliate Stentor coeruleus. We acquired and maintained the correct organism in laboratory culture and used micromanipulation and video microscopy to confirm Jennings’ observations. Despite significant individual variation, not addressed by Jennings, S. roeseli exhibits avoidance behaviors in a characteristic hierarchy of bending, ciliary alteration, contractions, and detachment, which is distinct from habituation or conditioning. Remarkably, the choice of contraction versus detachment is consistent with a fair coin toss. Such behavioral complexity may have had an evolutionary advantage in protist ecosystems, and the ciliate cortex may have provided mechanisms for implementing such behavior prior to the emergence of multicellularity. Our work resurrects Jennings’ pioneering insights and adds to the list of exceptional features, including regeneration [ 24 ], genome rearrangement [ 25 ], codon reassignment [ 26 ], and cortical inheritance [ 27 ], for which the ciliate clade is renowned.

Excitation-contraction coupling in smooth muscle is mediated by the Ca2+ - and calmodulin-dependent regulation of myosin light chain kinase. The precise mechanism of this regulation remains controversial, and several mathematical models have been proposed for the interaction of the three species. These models have previously been analyzed at steady state primarily by numerical simulation of differential equations, for which parameter values must be estimated from data. Here, we use the linear framework for timescale separation to demonstrate that models of this general kind can be solved analytically for an equilibrium steady state, without having to determine parameter values. This analysis leads to parameter-independent methods for discriminating between the models, for which we propose experiments that could be performed with existing methods.

Classification of texts by genre is an important application of natural language processing to literary corpora but remains understudied for premodern and non-English traditions. We develop a stylometric feature set for ancient Greek that enables identification of texts as prose or verse. The set contains over 20 primarily syntactic features, which are calculated according to custom, language-specific heuristics. Using these features, we classify almost all surviving classical Greek literature as prose or verse with >97% accuracy and F1 score, and further classify a selection of the verse texts into the traditional genres of epic and drama.

Computational stylometry has become an increasingly important aspect of literary criticism, but many humanists lack the technical expertise or language-specific NLP resources required to exploit computational methods. We demonstrate a stylometry toolkit for analysis of Latin literary texts, which is freely available at www.qcrit.org/stylometry. Our toolkit generates data for a diverse range of literary features and has an intuitive point-and-click interface. The features included have proven effective for multiple literary studies and are calculated using custom heuristics without the need for syntactic parsing. As such, the toolkit models one approach to the user-friendly generation of stylometric data, which could be extended to other premodern and non-English languages underserved by standard NLP resources.

Excitation-contraction coupling in smooth muscle is mediated by the Ca

Monoallelic point mutations in the gene encoding the cytosolic, NADP(+)-dependent enzyme isocitrate dehydrogenase 1 (IDH1) cause increased production of the oncometabolite 2-hydroxyglutarate (2-HG) in multiple cancers. Most IDH1 mutant tumors retain one wildtype (WT) IDH1 allele. Several studies have proposed that retention of this WT allele is protumorigenic by facilitating substrate channeling through a WT–mutant IDH1 heterodimer, with the WT subunit generating a local supply of α-ketoglutarate and NADPH that is then consumed by the mutant subunit to produce 2-HG. Here, we confirmed that coexpression of WT and mutant IDH1 subunits leads to formation of WT–mutant hetero-oligomers and increases 2-HG production. An analysis of a recently reported crystal structure of the WT–R132H IDH1 heterodimer and of in vitro kinetic parameters for 2-HG production, however, indicated that substrate channeling between the subunits is biophysically implausible. We also found that putative carbon-substrate flux between WT and mutant IDH1 subunits is inconsistent with the results of isotope tracing experiments in cancer cells harboring an endogenous monoallelic IDH1 mutation. Finally, using a mathematical model of WT–mutant IDH1 heterodimers, we estimated that the NADPH:NADP(+) ratio is higher in the cytosol than in the mitochondria, suggesting that NADPH is unlikely to be limiting for 2-HG production in the cytosol. These findings argue against supply of either substrate being limiting for 2-HG production by a cytosolic IDH1 mutant and suggest that the retention of a WT allele in IDH1 mutant tumors is not due to a requirement for carbon or cofactor flux between WT and mutant IDH1.

Experimental and theoretical studies have suggested that bifunctional enzymes catalyzing opposing modification and demodification reactions can confer steady-state concentration robustness to their substrates. However, the types of robustness and the biochemical basis for them have remained elusive. Here we report a systematic study of the most general biochemical reaction network for a bifunctional enzyme acting on a substrate with one modification site, along with eleven sub-networks with more specialized biochemical assumptions. We exploit ideas from computational algebraic geometry, introduced in previous work, to find a polynomial expression (an invariant) between the steady state concentrations of the modified and unmodified substrate for each network. We use these invariants to identify five classes of robust behavior: robust upper bounds on concentration, robust two-sided bounds on concentration ratio, hybrid robustness, absolute concentration robustness (ACR), and robust concentration ratio. This analysis demonstrates that robustness can take a variety of forms and that the type of robustness is sensitive to many biochemical details, with small changes in biochemistry leading to very different steady-state behaviors. In particular, we find that the widely-studied ACR requires highly specialized assumptions in addition to bifunctionality. An unexpected result is that the robust bounds derived from invariants are strictly tighter than those derived by ad hoc manipulation of the underlying differential equations, confirming the value of invariants as a tool to gain insight into biochemical reaction networks. Furthermore, invariants yield multiple experimentally testable predictions and illuminate new strategies for inferring enzymatic mechanisms from steady-state measurements.

This article examines a case study in Silius Italicus’ Punica using two distinct but complementary approaches to Flavian epic intertextuality: a methodological move to expand and further incorporate computational tools within philology, and a literary theoretical move to combine intertextuality and thematic interpretation. The case study focuses on the debates in the Carthaginian senate described in Punica 2 and 11, both of which Silius adapts from similar scenes in Livy while also drawing on Vergil’s Aeneid. Part 1 of the essay introduces a new tool for finding a range of inexact verbal parallels based on a bioinformatics technique known as sequence alignment. After comparing the method with two other computational tools, Diogenes and Tesserae, we assess our tool’s ability to detect intertexts in the Punica already noted in traditional scholarship. We then analyse a series of computationally identified parallels that have not been commented on previously and find that all three tools can reveal morphologically and syntactically similar phrases of apparent literary interest. Part 2 focuses on a feature of Silius’ triangulation of Livy and Vergil, the characterisation of the Carthaginian senator Hanno. Through allusions to Vergil’s Drances, Silius turns Hanno from a shrewd judge of Roman character and strength, as he appears in Livy, into a far more ambivalent, Quisling-like figure. Moreover, the effect of blending the two sources is to make more porous the distinctions between nationalities and other categories that structure the reader’s response to Hanno and to the Punica as a whole. In concluding, we suggest that the context in which these literary interactions take place - diplomacy and debate - itself figures the kind of negotiation taking place at a textual level between the various works and their worldviews. The conclusion unifies the methodological and theoretical parts of the essay under the rubric of “triangulation”, in part by drawing on the application of the term in the philosophy of Donald Davidson.

This paper examines a loose, popularized translation of Aeneid 1 and 4 published in 1870 by a small New England newspaper. Through evaluation of the translator’s preface and related material, I argue that the text was intended to be a response to the marginalization of the classics in school curricula and a useful pedagogical tool for attracting students to Vergil. I then consider the translation in relation to the long tradition of Vergilian travesty and propose that it can be read as a combinatorial parody that satirizes both the original epic and nineteenth-century American society in an attempt to increase interest in the classics.

Planarians are an important model organism for regeneration and stem cell research. A complete understanding of stem cell and regeneration dynamics in these animals requires time-lapse imaging in vivo, which has been difficult to achieve due to a lack of tissue-specific markers and the strong negative phototaxis of planarians. We have developed the Planarian Immobilization Chip (PIC) for rapid, stable immobilization of planarians for in vivo imaging without injury or biochemical alteration. The chip is easy and inexpensive to fabricate, and worms can be mounted for and removed after imaging within minutes. We show that the PIC enables significantly higher-stability immobilization than can be achieved with standard techniques, allowing for imaging of planarians at sub-cellular resolution in vivo using brightfield and fluorescence microscopy. We validate the performance of the PIC by performing time-lapse imaging of planarian wound closure and sequential imaging over days of head regeneration. We further show that the device can be used to immobilize Hydra, another photophobic regenerative model organism. The simple fabrication, low cost, ease of use, and enhanced specimen stability of the PIC should enable its broad application to in vivo studies of stem cell and regeneration dynamics in planarians and Hydra.

An important goal of systems biology is to develop quantitative models that explain how specific molecular features give rise to systems-level properties. Metabolic and regulatory pathways that contain multifunctional proteins are especially interesting to study from this perspective because they have frequently been observed to exhibit robustness: the ability for a system to perform its proper function even as levels of its components change. In this study, we use extensive biochemical data and algebraic modeling to develop and analyze a model that shows how robust behavior arises in the isocitrate dehydrogenase (IDH) regulatory system of Escherichia coli, which was shown in 1985 to experimentally exhibit robustness. E. coli IDH is regulated by reversible phosphorylation catalyzed by the bifunctional isocitrate dehydrogenase kinase/phosphatase (IDHKP), and the level of IDH activity determines whether carbon flux is directed through the glyoxylate bypass (for growth on two-carbon substrates) or the full tricarboxylic acid cycle. Our model, which incorporates recent structural data on IDHKP, identifies several specific biochemical features of the system (including homodimerization of IDH and bifunctionality of IDHKP) that provide a potential explanation for robustness. Using algebraic techniques, we derive an invariant that summarizes the steady-state relationship between the phospho-forms of IDH. We use the invariant in combination with kinetic data on IDHKP to calculate IDH activity at a range of total IDH levels and find that our model predicts robustness. Our work unifies much of the known biochemistry of the IDH regulatory system into a single quantitative framework and highlights the importance of constructing biochemically realistic models in systems biology.

Monoallelic point mutations in the cytosolic, NADP+-dependent enzyme isocitrate dehydrogenase 1 (IDH1) are associated with elevated production of the oncometabolite 2-hydroxyglutarate (2-HG) in multiple cancers. Production of 2-HG by mutant R132H IDH1 requires activity of a retained wild-type (WT) allele, suggesting possible channeling of α-ketoglutarate and NADPH in WT/R132H IDH1 heterodimers [1]. We report a mechanistic characterization of IDH1 2-HG production using a combination of mathematical modeling, in vivo kinetic measurements, and diffusion calculations. Numerical simulation of a biochemically realistic mathematical model of the IDH1 heterodimer indicated that substrate channeling would have a large impact on the dynamics of 2-HG generation. We then examined conversion of 13C labeled glucose and 13C labeled glutamine into labeled 2-HG in HT1080 cells and found that the dynamics are nearly identical in both cases. This result provides strong evidence against channeling because the production of 2-HG from glucose, but not glutamine, is dependent on WT IDH1. Calculations of substrate diffusion between the WT and mutant active sites of both isolated and agglomerated IDH1 heterodimers further support the absence of channeling. We propose instead that 2-HG production is sensitive to the global availability of substrates in the appropriate intracellular compartment, consistent with prior observations that 2-HG production by structurally analogous mutants of the mitochondrial isoform IDH2 is not WT-dependent [1]. Building on this observation, we report a strategy for inferring compartment-specific NADPH/NADP+ ratios using our model and steady-state measurements of 2-HG production by native IDH1 and by IDH1 artificially targeted to the mitochondrion.[1] Ward, P.S., C. Lu, J.R. Cross, O. Abdel-Wahab, R.L. Levine, G.K. Schwartz, and C.B. Thompson. 2013. J. Biol. Chem. 288: 3804-3815

Improving methods for high-throughput combinatorial chemistry has emerged as a major area of research because of the importance of rapidly synthesizing large numbers of chemical compounds for drug discovery and other applications. In this investigation, a novel microfluidic chip for performing parallel combinatorial chemical synthesis was developed. Unlike past microfluidic systems designed for parallel combinatorial chemistry, the chip is a single-layer device made of poly(dimethylsiloxane) that is extremely easy and inexpensive to fabricate. Using the chip, a 2×2 combinatorial series of amide-formation reactions was performed. The results of this combinatorial synthesis indicate that the new device is an effective platform for running parallel organic syntheses at significantly higher throughput than with past methodologies. Additionally, a design algorithm for scaling up the 2×2 combinatorial synthesis chip to address more complex cases was developed.

Background The yeast Saccharomyces cerevisiae relies on the high-osmolarity glycerol (HOG) signaling pathway to respond to increases in external osmolarity. The HOG pathway is rapidly activated under conditions of elevated osmolarity and regulates transcriptional and metabolic changes within the cell. Under normal growth conditions, however, a three-component phospho-relay consisting of the histidine kinase Sln1, the transfer protein Ypd1, and the response regulator Ssk1 represses HOG pathway activity by phosphorylation of Ssk1. This inhibition of the HOG pathway is essential for cellular fitness in normal osmolarity. Nevertheless, the extent to and mechanisms by which inhibition is robust to fluctuations in the concentrations of the phospho-relay components has received little attention. Results We established that the Sln1-Ypd1-Ssk1 phospho-relay is robust—it is able to maintain inhibition of the HOG pathway even after significant changes in the levels of its three components. We then developed a biochemically realistic mathematical model of the phospho-relay, which suggested that robustness is due to buffering by a large excess pool of Ypd1. We confirmed experimentally that depletion of the Ypd1 pool results in inappropriate activation of the HOG pathway. Conclusions We identified buffering by an intermediate component in excess as a novel mechanism through which a phospho-relay can achieve robustness. This buffering requires multiple components and is therefore unavailable to two-component systems, suggesting one important advantage of multi-component relays. Electronic supplementary material The online version of this article (doi:10.1186/s12918-015-0158-y) contains supplementary material, which is available to authorized users.