bluuebunny/crossref_metadata_2025_split huggingface dataset:
Spontaneous Alignment of Myotubes through Myogenic Progenitor Cell Migration
Authors: Mehanna Lauren E. , Osborne Adrianna R. , Peterson Charlotte A. , Berron Brad J. 
Published: July 2023

AbstractIn large volume muscle injuries, widespread damage to muscle fibers and the surrounding connective tissue prevents myogenic progenitor cells (MPCs) from initiating repair. There is a clinical need to rapidly fabricate large muscle tissue constructs for integration at the site of large volume muscle injuries. Most strategies for myotube alignment require microfabricated structures or prolonged orientation times. We utilize the MPC’s natural propensity to close gaps across an injury site to guide alignment on collagen I. When MPCs are exposed to an open boundary free of cells, they migrate unidirectionally into the cell-free region and align perpendicular to the original boundary direction. We study the utility of this phenomenon with biotin - streptavidin adhesion to position the cells on the substrate, and then demonstrate the robustness of this strategy with unmodified cells, creating a promising tool for MPC patterning without interrupting their natural function. We pre-position MPCs in straight-line patterns separated with small gaps. This temporary positioning initiates the migratory nature of the MPCs to align and form myotubes across the gaps, similar to how they migrate and align with a single open boundary. There is a directional component to the MPC migration perpendicular (90°) to the original biotin-streptavidin surface patterns. The expression of myosin heavy chain, the motor protein of muscle thick filaments, is confirmed through immunocytochemistry (ICC) in myotubes generated from MPCs in our patterning process, acting as a marker of skeletal muscle differentiation. The rapid and highly specific binding of biotin-streptavidin allows for quick formation of temporary patterns, with MPC alignment based on natural regenerative behavior rather than complex fabrication techniques.Impact StatementPositioning myogenic progenitor cells (MPCs) into straight-line patterns with intentional spacings initiates the migration of these cells to bridge these gaps, mimicking their behavior in response to small-scale injuries. By creating repetitions of patterned cells and spacings, we have demonstrated rapid migration and alignment of MPCs, which differentiate into a long-range 2D layer of aligned myotubes.
Burkholderia pseudomalleiBicA protein promotes pathogenicity in macrophages by regulating Invasion, Intracellular Survival, and Virulence
Authors: Stockton Jacob L , Khakhum Nittaya , Stevenson Heather L. , Torres Alfredo G 
Published: July 2023

AbstractBurkholderia pseudomallei (Bpm)is the causative agent of melioidosis disease.Bpmis a facultative intracellular pathogen with a complex lifecycle inside host cells. Pathogenic success depends on a variety of virulence factors with one of the most critical being the type 6 secretion system (T6SS).Bpmuses the T6SS to move into neighboring cells, resulting in multinucleated giant cells (MNGCs) formation, a strategy used to disseminate from cell-to-cell. Our prior study using a dual RNA-seq analysis to dissect T6SS-mediated virulence on intestinal epithelial cells identified BicA as a factor upregulated in a T6SS mutant (1). BicA regulates both type 3 secretion system (T3SS) and T6SSs; however, the extent of its involvement during disease progression is unclear. To fully dissect the role of BicA during systemic infection, we used two macrophage cell lines paired with a pulmonaryin vivochallenge murine model. We found that ΔbicAhas a distinct intracellular replication defect in both immortalized and primary macrophages that begins as early as 1 h post-infection. This intracellular defect is linked with the lack of cell-to-cell dissemination and MNGC formation, as well as a defect on T3SS expression. Thein vitrophenotype translatedin vivoas ΔbicAwas attenuated in a pulmonary model of infection; demonstrating a distinct macrophage activation profile and lack of pathological features present in the wild type. Overall, these results highlight the role of BicA in regulating intracellular virulence and demonstrate that specific regulation of secretion systems has a significant effect on host response andBpmpathogenesis.ImportanceMelioidosis is an understudied tropical disease that still results in ∼50% fatalities from those infected patients. It is caused by the Gram-negative bacillusBurkholderia pseudomallei(Bpm). Bpmis an intracellular pathogen that disseminates from the infected cell to target organs, causing disseminated disease. Regulation of secretion systems involved in entry and cell-to-cell spread is poorly understood. In this work, we characterize the role of BicA as a regulator of secretion systems during infection of macrophagesin vitroandin vivo. Understanding how these virulence factors are controlled will help us determine their influence on the host cells and define the macrophage responses associated with bacterial clearance.
ZIF-1-mediated degradation of endogenous and heterologous zinc finger proteins in theC. elegansgerm line
Authors: Schwartz Aaron Z.A. , Abdu Yusuff , Nance Jeremy 
Published: July 2023

AbstractRapid and conditional protein depletion is the gold standard genetic tool for deciphering the molecular basis of developmental processes. Previously, we showed that by conditionally expressing the E3 ligase substrate adaptor ZIF-1 inCaenorhabditis eleganssomatic cells, proteins tagged with the first CCCH Zn finger (ZF1) domain from the germline regulator PIE-1 degrade rapidly, resulting in loss-of-function phenotypes. The described role of ZIF-1 is to clear PIE-1 and several other CCCH Zn finger proteins from early somatic cells, helping to enrich them in germline precursor cells. Here, we show that proteins tagged with the PIE-1 ZF1 domain are subsequently cleared from primordial germ cells in embryos and from undifferentiated germ cells in larvae and adults by ZIF-1. We harness germline ZIF-1 activity to degrade a ZF1-tagged heterologous protein from PGCs and show that its depletion produces phenotypes equivalent to those of a null mutation. Our findings reveal that ZIF-1 switches roles from degrading CCCH Zn finger proteins in somatic cells to clearing them from undifferentiated germ cells, and that ZIF-1 activity can be harnessed as a new genetic tool to study the early germ line.
DUX4 is a common driver of immune evasion and immunotherapy failure in metastatic cancers
Authors: Pineda Jose Mario Bello , Bradley Robert K. 
Published: July 2023

ABSTRACTCancer immune evasion contributes to checkpoint immunotherapy failure in many patients with metastatic cancers. The embryonic transcription factor DUX4 was recently characterized as a suppressor of interferon-γ signaling and antigen presentation that is aberrantly expressed in a small subset of primary tumors. Here, we report thatDUX4expression is a common feature of metastatic tumors, with ∼10-50% of advanced bladder, breast, kidney, prostate, and skin cancers expressingDUX4.DUX4expression is significantly associated with immune cell exclusion and decreased objective response to PD-L1 blockade in a large cohort of urothelial carcinoma patients.DUX4expression is a significant predictor of survival even after accounting for tumor mutational burden and other molecular and clinical features in this cohort, withDUX4expression associated with a median reduction in survival of over one year. Our data motivate future attempts to develop DUX4 as a biomarker and therapeutic target for checkpoint immunotherapy resistance.
A temperature-sensitive metabolic valve and a transcriptional feedback loop drive rapid homeoviscous adaptation inEscherichia coli
Authors: Hoogerland Loles , van den Berg Stefan , Zoumaro-Djayoon Adja , Geurken Esther , Yang Flora , Bruggeman Frank , Bokinsky Gregory 
Published: July 2023

AbstractAll free-living microorganisms homeostatically maintain the fluidity of their membranes by adapting lipid composition to environmental temperatures. A quantitative description of how organisms maintain constant fluidity at all growth temperatures has not been achieved. By quantifying both enzymes and metabolic intermediates of theEscherichia colifatty acid and phospholipid synthesis pathways, we discover howE. colimeasures steady-state temperature and restores optimal membrane fluidity within a single generation after temperature shocks. The first element of the system is a temperature-sensitive metabolic valve that allocates flux between the saturated and unsaturated fatty acid synthesis pathways. The second element is a transcription-based negative feedback loop that counteracts the temperature-sensitive valve. The combination of these elements accelerates membrane adaptation by causing a transient overshoot in the synthesis of saturated or unsaturated fatty acids following temperature shocks. This overshoot strategy accelerates membrane adaptation, and is comparable to increasing the temperature of a water bath by adding water that is excessively hot rather than adding water at the desired temperature. These properties are captured in a quantitative model, which we further use to show how hard-wired parameters calibrate the system to generate membrane compositions that maintain constant fluidity across a wide range of temperatures. We hypothesize that core design features of theE. colisystem will prove to be ubiquitous features of homeoviscous adaptation systems.
Proteome-wide copy-number estimation from transcriptomics
Authors: Sweatt Andrew J. , Griffiths Cameron D. , Paudel B. Bishal , Janes Kevin A. 
Published: July 2023

AbstractProtein copy numbers constrain systems-level properties of regulatory networks, but absolute proteomic data remain scarce compared to transcriptomics obtained by RNA sequencing. We addressed this persistent gap by relating mRNA to protein statistically using best-available data from quantitative proteomics–transcriptomics for 4366 genes in 369 cell lines. The approach starts with a central estimate of protein copy number and hierarchically appends mRNA-protein and mRNA-mRNA dependencies to define an optimal gene-specific model that links mRNAs to protein. For dozens of independent cell lines and primary prostate samples, these protein inferences from mRNA outmatch stringent null models, a count-based protein-abundance repository, and empirical protein-to-mRNA ratios. The optimal mRNA-to-protein relationships capture biological processes along with hundreds of known protein-protein interaction complexes, suggesting mechanistic relationships are embedded. We use the method to estimate viral-receptor abundances of CD55–CXADR from human heart transcriptomes and build 1489 systems-biology models of coxsackievirus B3 infection susceptibility. When applied to 796 RNA sequencing profiles of breast cancer from The Cancer Genome Atlas, inferred copy-number estimates collectively reclassify 26% of Luminal A and 29% of Luminal B tumors. Protein-based reassignments strongly involve a pharmacologic target for luminal breast cancer (CDK4) and an α-catenin that is often undetectable at the mRNA level (CTTNA2). Thus, by adopting a gene-centered perspective of mRNA-protein covariation across different biological contexts, we achieve accuracies comparable to the technical reproducibility limits of contemporary proteomics. The collection of gene-specific models is assembled as a web tool for users seeking mRNA-guided predictions of absolute protein abundance (http://janeslab.shinyapps.io/Pinferna).Significance statementIt is easier to quantify mRNA in cells than it is to quantify protein, but proteins are what execute most cellular functions. Even though protein is synthesized from mRNA in cells, relating a cellular quantity of mRNA to a quantity of protein is challenging. Here, we bring together quantitative measures of mRNA and protein for 4366 genes in 369 different cultured cell types to build equations that predict protein abundance from the abundance of mRNAs expressed. These equations capture facets of biological regulation and work better than existing alternatives that rely on consensus values or ratios. Since mRNA measurements are more widespread than protein, this study makes new analyses possible by protein estimation from mRNA.
Calcineurin is required forCandida glabrataPdr1 transcriptional activation
Authors: Vu Bao Gia , Simonicova Lucia , Moye-Rowley W. Scott 
Published: July 2023

AbstractFluconazole is the most commonly used antifungal today. A result of this has been the inevitable selection of fluconazole resistant organisms. This is an especially acute problem in the pathogenic yeastCandida glabrata. Elevated minimal inhibitory concentrations (MICs) for fluconazole inC. glabrataare frequently associated with substitution mutations within the Zn2Cys6 zinc cluster-containing transcription factor-encoding genePDR1. These mutant Pdr1 regulators drive constitutively high expression of target genes likeCDR1that encodes an ATP-binding cassette transporter thought to act as a drug efflux pump. Exposure ofC. glabratato fluconazole induced expression of both Pdr1 andCDR1, although little is known of the molecular basis underlying the upstream signals that trigger Pdr1 activation. Here, we show that the protein phosphatase calcineurin is required for fluconazole-dependent induction of Pdr1 transcriptional regulation. Calcineurin catalytic activity is required for normal Pdr1 regulation and a hyperactive form of this phosphatase can increase resistance to the echinocandin caspofungin but does not show a similar elevation for fluconazole resistance. Loss of calcineurin from strains expressing two different gain-of-function forms of Pdr1 also caused a decrease inCDR1expression and fluconazole resistance, demonstrating that even these hyperactive Pdr1 regulatory mutants cannot bypass the requirement for calcineurin. Our data implicate calcineurin activity as a link tying azole and echinocandin resistance together via the control of transcription factor activity.ImportanceWhile drug resistant microorganisms are a problem in treatment of all infectious disease, this is an especially acute problem with fungi due to the existence of only 3 classes of antifungal drugs, including the azole drug fluconazole. In the pathogenic yeastCandida glabrata, mutant forms of a transcription factor called Pdr1 are commonly associated with fluconazole resistance and poor clinical outcomes. Here we identify a protein phosphatase called calcineurin that is required for fluconazole-dependent induction of Pdr1 transcriptional activation and associated drug resistance. Gain-of-function mutant forms of Pdr1 still required the presence of calcineurin to confer normally elevated fluconazole resistance. Previous studies showed that calcineurin is required for resistance to the echinocandin class of antifungal drugs and our data demonstrate this protein phosphatase is also required for azole drug resistance. Calcineurin plays a central role in resistance to two of the three major classes of antifungal drugs inC. glabrata.
Scaling, correlations, and artifacts in biological timeseries
Authors: Faerberg Denis F. , Gurarie Victor , Ruvinsky Ilya 
Published: July 2023

SummaryTime is a key variable in biology. Conceptual and methodological advances focused recent studies of timing in biological systems on monitoring trait dynamics in singled individuals. Here we contribute to this effort by analyzing general properties of individual timeseries. We make four broad claims. First, we show that for traits that range from behavior in animals, to growth in plants, to division timing in cell lineages, faster timeseries tend to be comprised of consistently shorter sub-stages, not a few unusually fast sub-stages. We demonstrate that this property constitutes a particular type of scaling that can be readily detected by a straightforward comparison of absolute and relative variability within timeseries data for any trait. We show that correlations within timeseries are necessary and sufficient for the type of scaling we describe and infer that the ubiquitous occurrence of scaling results from natural correlations within the continuous processes that govern trait dynamics. Second, failing to observe scaling indicates a break in correlation and, we argue, a switch in the mechanisms underlying the rate of trait change. Third, we caution that analyses of individual timeseries are highly susceptible to batch effects, but contend that it is possible to detect signal despite the artifactual correlations introduced by batching. Finally, we show that the timing of biological process that scale is overdispersed compared to matched but uncorrelated comparisons; this is an advantage in variable environments. Our results advance understanding of common properties of biological timeseries and offer practical tools for their further study.
Integrative reappraisal of the Amazonian nurse frogAllobates gasconi(Morales 2002) based on topotypical data, with implications for the systematics and taxonomy of a large species complex
Authors: Ferreira Anthony S. , Ferrão Miquéias , Cunha-Machado Antonio S. , Magnusson William E. , Hanken James , Lima Albertina P. 
Published: July 2023

AbstractTaxonomic uncertainty at the species level compromises our knowledge of biodiversity, conservation and systematics. The impact of such uncertainty is heightened in megadiverse regions such as Amazonia due to high levels of cryptic diversity. We used integrative taxonomy based on newly collected topotypical specimens to redescribe the Amazonian nurse frogAllobates gasconiand infer its phylogenetic relationships. This species was described in 2002 based solely on morphology, but several characters crucial for the reliable diagnosis of species inAllobateswere not considered. Our results show thatA. gasconisensu stricto is not a member of theA. caeruleodactylusclade as previously claimed, but is a member of theA. trilineatusclade.Allobates gasconiis readily distinguished from congeners by a combination of morphological and bioacoustic characters; a revised diagnosis is provided. The type series ofA. gasconicomprises more than one species, and we exclude a paratype from lower Juruá River. The species is restricted geographically to flooded environments in the middle and upper Juruá River in Brazil and in the Ucayali River in Peru. The initial misidentification, subsequent absence of topotypic molecular and acoustic data, and the poor preservation condition of the type series have contributed to taxonomic confusion sinceA. gasconiwas first described. The descriptions of other species ofAllobatespublished more than two decades ago were based mainly on gross morphology and we recommend integrative taxonomic revisions to elucidate their systematics.
Atomic-level architecture ofCaulobacter crescentusflagellar filaments provide evidence for multi-flagellin filament stabilization
Authors: Sanchez Juan C. , Montemayor Eric J. , Ploscariu Nicoleta T. , Parrell Daniel , Baumgardt Joseph K. , Yang Jie E. , Sibert Bryan , Cai Kai , Wright Elizabeth R. 
Published: July 2023

AbstractFlagella are dynamic, ion-powered machines with assembly pathways that are optimized for efficient flagella production. In bacteria, dozens of genes are coordinated at specific times in the cell lifecycle to generate each component of the flagellum. This is the case forCaulobacter crescentus, but little is known about why this species encodes six different flagellin genes. Furthermore, little is known about the benefits multi-flagellin species possess over single flagellin species, if any, or what molecular properties allow for multi-flagellin filaments to assemble. Here we present an in-depth analysis of several single flagellin filaments fromC. crescentus,including an extremely well-resolved structure of a bacterial flagellar filament. We highlight key molecular interactions that differ between each bacterial strain and speculate how these interactions may alleviate or impose helical strain on the overall architecture of the filament. We detail conserved residues within the flagellin subunit that allow for the synthesis of multi-flagellin filaments. We further comment on how these molecular differences impact bacterial motility and highlight how no single flagellin filament achieves wild-type levels of motility, suggestingC. crescentushas evolved to produce a filament optimized for motility comprised of six flagellins. Finally, we highlight an ordered arrangement of glycosylation sites on the surface of the filaments and speculate how these sites may protect the β-hairpin located on the surface exposed domain of the flagellin subunit.
Altered GM1 catabolism affects NMDAR-mediated Ca2+signaling at ER-PM junctions and increases synaptic spine formation
Authors: Weesner Jason A. , Annunziata Ida , van de Vlekkert Diantha , Robinson Camenzind G. , Campos Yvan , Mishra Ashutosh , Fremuth Leigh E. , Gomero Elida , Hu Huimin , d’Azzo Alessandra 
Published: July 2023

SummaryEndoplasmic reticulum–plasma membrane (ER-PM) junctions mediate Ca2+flux across neuronal membranes. The properties of these membrane contact sites are defined by their lipid content, but little attention has been given to glycosphingolipids (GSLs). Here, we show that GM1-ganglioside, an abundant GSL in neuronal membranes, is integral to ER-PM junctions; it interacts with synaptic proteins/receptors and regulates Ca2+signaling. In a model of the neurodegenerative lysosomal storage disease, GM1-gangliosidosis, pathogenic accumulation of GM1 at ER-PM junctions due to β-galactosidase deficiency drastically alters neuronal Ca2+homeostasis. Mechanistically, we show that GM1 interacts with the phosphorylated NMDAR Ca2+channel, thereby increasing Ca2+flux, activating ERK signaling, and increasing the number of synaptic spines without increasing synaptic connectivity. Thus, GM1 clustering at ER-PM junctions alters synaptic plasticity and exacerbates the generalized neuronal cell death characteristic of GM1-gangliosidosis.
Position-Specific Enrichment Ratio Matrix scores predict antibody variant properties from deep sequencing data
Authors: Smith Matthew D. , Case Marshall A. , Makowski Emily K. , Tessier Peter M. 
Published: July 2023

AbstractMotivationDeep sequencing of antibody and related protein libraries after phage or yeast-surface display sorting is widely used to identify variants with increased affinity, specificity and/or improvements in key biophysical properties. Conventional approaches for identifying optimal variants typically use the frequencies of observation in enriched libraries or the corresponding enrichment ratios. However, these approaches disregard the vast majority of deep sequencing data and often fail to identify the best variants in the libraries.ResultsHere, we present a method, Position-Specific Enrichment Ratio Matrix (PSERM) scoring, that uses entire deep sequencing datasets from pre- and post-selections to score each observed protein variant. The PSERM scores are the sum of the site-specific enrichment ratios observed at each mutated position. We find that PSERM scores are much more reproducible and correlate more strongly with experimentally measured properties than frequencies or enrichment ratios, including for multiple antibody properties (affinity and non-specific binding) for a clinical-stage antibody (emibetuzumab). We expect that this method will be broadly applicable to diverse protein engineering campaigns.AvailabilityAll deep sequencing datasets and code to do the analyses presented within are available via GitHub.ContactPeter Tessier,ptessier@umich.eduSupplementary informationSupplementary data are available at Bioinformatics online.