Microscopic biophysical model of self-organization in tissue due to feedback between cell- and macroscopic-scale forces

We develop a microscopic biophysical model for self-organization and reshaping of artificial tissue, that is codriven by microscopic active forces between cells and an extracellular matrix (ECM), and macroscopic forces that develop within the tissue, finding close agreement with experiment. Microscopic active forces are stimulated by μm-scale interactions between cells and the ECM within which they exist, and when large numbers of cells act together these forces drive, and are affected by, macroscopic-scale self-organization and reshaping of tissues in a feedback loop. To understand this loop, there is a need to (1) construct microscopic biophysical models that can simulate these processes for the very large number of cells found in tissues, (2) validate and calibrate those models against experimental data, and (3) understand the active feedback between cells and the extracellular matrix, and its relationship to macroscopic self-organization and reshaping of tissue. Our microscopic biophysical model consists of a contractile network representing the ECM, that interacts with a large number of cells via dipole forces, to describe macroscopic self-organization and reshaping of tissue. We solve the model using simulated annealing, finding close agreement with experiments on artificial neural tissue. We discuss the calibration of model parameters. We conclude that feedback between microscopic cell-ECM dipole interactions and tissue-scale forces is a key factor in driving macroscopic self-organization and reshaping of tissue. We discuss the application of the biophysical model to the simulation and rational design of artificial tissues

The concentration of three anti-seizure medications in hair: the effects of hair color, controlling for dose and age

BACKGROUND: This paper assess the relationship between the quantity of three anti-seizure medications in hair and the color of the analyzed hair, while controlling for the effects of dose, dose duration, and patient age for 140 clinical patients undergoing anti-seizure therapy. Three drugs are assessed: carbamazepine (40 patients), valproic acid (40 patients), and phenytoin (60 patients). The relationship between hair assay results, hair color, dose, dose duration, and age is modeled using an analysis of covariance. The covariance model posits the hair assay results as the dependent variable, the hair color as the qualitative categorical independent variable, and dose, dose duration, and age as covariates. The null hypothesis assessed is that there is a no relationship between hair color and the quantity of analyte determined by hair assay such that darker colored hair will demonstrate higher concentrations of analyte than lighter colored hair. RESULTS: The analysis reveals that there is a significant relationship between dose and concentration for all hair color categories independent of the other covariates or the categorical independent variable. CONCLUSION: There does not appear to be any relationship between carbamazepine concentration and hair color. There is a weak relationship between hair color and valproic acid concentration, which the data suggest may be mediated by age. There is a significant, moderate relationship between phenytoin concentration and hair color such that darker colored hair has greater concentration values than lighter colored hair

Genome-wide maps of alkylation damage, repair, and mutagenesis in yeast reveal mechanisms of mutational heterogeneity

DNA base damage is an important contributor to genome instability, but how the formation and repair of these lesions is affected by the genomic landscape and contributes to mutagenesis is unknown. Here, we describe genome-wide maps of DNA base damage, repair, and mutagenesis at single nucleotide resolution in yeast treated with the alkylating agent methyl methanesulfonate (MMS). Analysis of these maps revealed that base excision repair (BER) of alkylation damage is significantly modulated by chromatin, with faster repair in nucleosome-depleted regions, and slower repair and higher mutation density within strongly positioned nucleosomes. Both the translational and rotational settings of lesions within nucleosomes significantly influence BER efficiency; moreover, this effect is asymmetric relative to the nucleosome dyad axis and is regulated by histone modifications. Our data also indicate that MMS-induced mutations at adenine nucleotides are significantly enriched on the nontranscribed strand (NTS) of yeast genes, particularly in BER-deficient strains, due to higher damage formation on the NTS and transcription-coupled repair of the transcribed strand (TS). These findings reveal the influence of chromatin on repair and mutagenesis of base lesions on a genome-wide scale and suggest a novel mechanism for transcription-associated mutation asymmetry, which is frequently observed in human cancers

Linearization of Cohomology-free Vector Fields

We study the cohomological equation for a smooth vector field on a compact manifold. We show that if the vector field is cohomology free, then it can be embedded continuously in a linear flow on an Abelian group

Global Analyses Of Ceratocystis Cacaofunesta Mitochondria: From Genome To Proteome.

The ascomycete fungus Ceratocystis cacaofunesta is the causal agent of wilt disease in cacao, which results in significant economic losses in the affected producing areas. Despite the economic importance of the Ceratocystis complex of species, no genomic data are available for any of its members. Given that mitochondria play important roles in fungal virulence and the susceptibility/resistance of fungi to fungicides, we performed the first functional analysis of this organelle in Ceratocystis using integrated omics approaches. The C. cacaofunesta mitochondrial genome (mtDNA) consists of a single, 103,147-bp circular molecule, making this the second largest mtDNA among the Sordariomycetes. Bioinformatics analysis revealed the presence of 15 conserved genes and 37 intronic open reading frames in C. cacaofunesta mtDNA. Here, we predicted the mitochondrial proteome (mtProt) of C. cacaofunesta, which is comprised of 1,124 polypeptides - 52 proteins that are mitochondrially encoded and 1,072 that are nuclearly encoded. Transcriptome analysis revealed 33 probable novel genes. Comparisons among the Gene Ontology results of the predicted mtProt of C. cacaofunesta, Neurospora crassa and Saccharomyces cerevisiae revealed no significant differences. Moreover, C. cacaofunesta mitochondria were isolated, and the mtProt was subjected to mass spectrometric analysis. The experimental proteome validated 27% of the predicted mtProt. Our results confirmed the existence of 110 hypothetical proteins and 7 novel proteins of which 83 and 1, respectively, had putative mitochondrial localization. The present study provides the first partial genomic analysis of a species of the Ceratocystis genus and the first predicted mitochondrial protein inventory of a phytopathogenic fungus. In addition to the known mitochondrial role in pathogenicity, our results demonstrated that the global function analysis of this organelle is similar in pathogenic and non-pathogenic fungi, suggesting that its relevance in the lifestyle of these organisms should be based on a small number of specific proteins and/or with respect to differential gene regulation. In this regard, particular interest should be directed towards mitochondrial proteins with unknown function and the novel protein that might be specific to this species. Further functional characterization of these proteins could enhance our understanding of the role of mitochondria in phytopathogenicity.149

Highly Recombinant VGII Cryptococcus gattii Population Develops Clonal Outbreak Clusters through both Sexual Macroevolution and Asexual Microevolution

ABSTRACTAn outbreak of the fungal pathogen Cryptococcus gattii began in the Pacific Northwest (PNW) in the late 1990s. This outbreak consists of three clonal subpopulations: VGIIa/major, VGIIb/minor, and VGIIc/novel. Both VGIIa and VGIIc are unique to the PNW and exhibit increased virulence. In this study, we sequenced the genomes of isolates from these three groups, as well as global isolates, and analyzed a total of 53 isolates. We found that VGIIa/b/c populations show evidence of clonal expansion in the PNW. Whole-genome sequencing provided evidence that VGIIb originated in Australia, while VGIIa may have originated in South America, and these were likely independently introduced. Additionally, the VGIIa outbreak lineage may have arisen from a less virulent clade that contained a mutation in the MSH2 ortholog, but this appears to have reverted in the VGIIa outbreak strains, suggesting that a transient mutator phenotype may have contributed to adaptation and evolution of virulence in the PNW outbreak. PNW outbreak isolates share genomic islands, both between the clonal lineages and with global isolates, indicative of sexual recombination. This suggests that VGII C.gattii has undergone sexual reproduction, either bisexual or unisexual, in multiple locales contributing to the production of novel, virulent subtypes. We also found that the genomes of two basal VGII isolates from HIV+ patients contain an introgression tract spanning three genes. Introgression substantially contributed to intra-VGII polymorphism and likely occurred through sexual reproduction with VGI. More broadly, these findings illustrate how both microevolution and sexual reproduction play central roles in the development of infectious outbreaks from avirulent or less virulent progenitors.IMPORTANCECryptococcus gattii is the causative agent responsible for ongoing infections in the Pacific Northwest of the United States and western Canada. The incidence of these infections increased dramatically in the 1990s and remains elevated. These infections are attributable to three clonal lineages of C.gattii, VGIIa, VGIIb, and VGIIc, with only VGIIa identified once previously in the Pacific Northwest prior to the start of the outbreak, albeit in a less virulent form. This study addresses the origin and emergence of this outbreak, using whole-genome sequencing and comparison of both outbreak and global isolates. We show that VGIIa arose mitotically from a less virulent clonal group, possibly via the action of a mutator phenotype, while VGIIb was likely introduced from Australia, and VGIIc appears to have emerged in the United States or in an undersampled locale via sexual reproduction. This work shows that multiple processes can contribute to the emergence of an outbreak

Genome And Secretome Analysis Of The Hemibiotrophic Fungal Pathogen, Moniliophthora Roreri, Which Causes Frosty Pod Rot Disease Of Cacao: Mechanisms Of The Biotrophic And Necrotrophic Phases

Background: The basidiomycete Moniliophthora roreri is the causal agent of Frosty pod rot (FPR) disease of cacao (Theobroma cacao), the source of chocolate, and FPR is one of the most destructive diseases of this important perennial crop in the Americas. This hemibiotroph infects only cacao pods and has an extended biotrophic phase lasting up to sixty days, culminating in plant necrosis and sporulation of the fungus without the formation of a basidiocarp.Results: We sequenced and assembled 52.3 Mb into 3,298 contigs that represent the M. roreri genome. Of the 17,920 predicted open reading frames (OFRs), 13,760 were validated by RNA-Seq. Using read count data from RNA sequencing of cacao pods at 30 and 60 days post infection, differential gene expression was estimated for the biotrophic and necrotrophic phases of this plant-pathogen interaction. The sequencing data were used to develop a genome based secretome for the infected pods. Of the 1,535 genes encoding putative secreted proteins, 1,355 were expressed in the biotrophic and necrotrophic phases. Analysis of the data revealed secretome gene expression that correlated with infection and intercellular growth in the biotrophic phase and invasive growth and plant cellular death in the necrotrophic phase.Conclusions: Genome sequencing and RNA-Seq was used to determine and validate the Moniliophthora roreri genome and secretome. High sequence identity between Moniliophthora roreri genes and Moniliophthora perniciosa genes supports the taxonomic relationship with Moniliophthora perniciosa and the relatedness of this fungus to other basidiomycetes. Analysis of RNA-Seq data from infected plant tissues revealed differentially expressed genes in the biotrophic and necrotrophic phases. 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Gene Copy-Number Variation in Haploid and Diploid Strains of the Yeast Saccharomyces cerevisiae

The increasing ability to sequence and compare multiple individual genomes within a species has highlighted the fact that copy-number variation (CNV) is a substantial and underappreciated source of genetic diversity. Chromosome-scale mutations occur at rates orders of magnitude higher than base substitutions, yet our understanding of the mechanisms leading to CNVs has been lagging. We examined CNV in a region of chromosome 5 (chr5) in haploid and diploid strains of Saccharomyces cerevisiae. We optimized a CNV detection assay based on a reporter cassette containing the SFA1 and CUP1 genes that confer gene dosage-dependent tolerance to formaldehyde and copper, respectively. This optimized reporter allowed the selection of low-order gene amplification events, going from one copy to two copies in haploids and from two to three copies in diploids. In haploid strains, most events involved tandem segmental duplications mediated by nonallelic homologous recombination between flanking direct repeats, primarily Ty1 elements. In diploids, most events involved the formation of a recurrent nonreciprocal translocation between a chr5 Ty1 element and another Ty1 repeat on chr13. In addition to amplification events, a subset of clones displaying elevated resistance to formaldehyde had point mutations within the SFA1 coding sequence. These mutations were all dominant and are proposed to result in hyperactive forms of the formaldehyde dehydrogenase enzyme

4-1BBL-containing leukemic extracellular vesicles promote immunosuppressive effector regulatory T cells

Chronic and acute myeloid leukemia evade immune system surveillance and induce immunosuppression by expanding proleukemic Foxp31 regulatory T cells (Tregs). High levels of immunosuppressive Tregs predict inferior response to chemotherapy, leukemia relapse, and shorter survival. However, mechanisms that promote Tregs in myeloid leukemias remain largely unexplored. Here, we identify leukemic extracellular vesicles (EVs) as drivers of effector proleukemic Tregs. Using mouse model of leukemia-like disease, we found that Rab27adependent secretion of leukemic EVs promoted leukemia engraftment, which was associated with higher abundance of activated, immunosuppressive Tregs. Leukemic EVs attenuated mTOR-S6 and activated STAT5 signaling, as well as evoked significant transcriptomic changes in Tregs. We further identified specific effector signature of Tregs promoted by leukemic EVs. Leukemic EVs-driven Tregs were characterized by elevated expression of effector/tumor Treg markers CD39, CCR8, CD30, TNFR2, CCR4, TIGIT, and IL21R and included 2 distinct effector Treg (eTreg) subsets: CD301CCR8hiTNFR2hi eTreg1 and CD391TIGIThi eTreg2. Finally, we showed that costimulatory ligand 4-1BBL/CD137L, shuttled by leukemic EVs, promoted suppressive activity and effector phenotype of Tregs by regulating expression of receptors such as CD30 and TNFR2. Collectively, our work highlights the role of leukemic extracellular vesicles in stimulation of immunosuppressive Tregs and leukemia growth. We postulate that targeting of Rab27a-dependent secretion of leukemic EVs may be a viable therapeutic approach in myeloid neoplasms

Analysis of a Food-Borne Fungal Pathogen Outbreak: Virulence and Genome of a Mucor circinelloides Isolate from Yogurt

ABSTRACTFood-borne pathogens are ongoing problems, and new pathogens are emerging. The impact of fungi, however, is largely underestimated. Recently, commercial yogurts contaminated with Mucor circinelloides were sold, and >200 consumers became ill with nausea, vomiting, and diarrhea. Mucoralean fungi cause the fatal fungal infection mucormycosis, whose incidence has been continuously increasing. In this study, we isolated an M.circinelloides strain from a yogurt container, and multilocus sequence typing identified the strain as Mucor circinelloides f. circinelloides. M. circinelloides f. circinelloides is the most virulent M.circinelloides subspecies and is commonly associated with human infections, whereas M.circinelloides f. lusitanicus and M.circinelloides f. griseocyanus are less common causes of infection. Whole-genome analysis of the yogurt isolate confirmed it as being close to the M. circinelloides f. circinelloides subgroup, with a higher percentage of divergence with the M.circinelloides f. lusitanicus subgroup. In mating assays, the yogurt isolate formed sexual zygospores with the (−) M. circinelloides f. circinelloides tester strain, which is congruent with its sex locus encoding SexP, the (+) mating type sex determinant. The yogurt isolate was virulent in murine and wax moth larva host systems. In a murine gastromucormycosis model, Mucor was recovered from fecal samples of infected mice for up to 10days, indicating that Mucor can survive transit through the GI tract. In interactions with human immune cells, M.circinelloides f. lusitanicus induced proinflammatory cytokines but M. circinelloides f. circinelloides did not, which may explain the different levels of virulence in mammalian hosts. This study demonstrates that M.circinelloides can spoil food products and cause gastrointestinal illness in consumers and may pose a particular risk to immunocompromised patients.IMPORTANCEThe U.S. FDA reported that yogurt products were contaminated with M. circinelloides, a mucoralean fungal pathogen, and >200 consumers complained of symptoms, including vomiting, nausea, and diarrhea. The manufacturer voluntarily withdrew the affected yogurt products from the market. Compared to other food-borne pathogens, including bacteria, viruses, and parasites, less focus has been placed on the risk of fungal pathogens. This study evaluates the potential risk from the food-borne fungal pathogen M. circinelloides that was isolated from the contaminated commercial yogurt. We successfully cultured an M.circinelloides isolate and found that the isolate belongs to the species M.circinelloides f. circinelloides, which is often associated with human infections. In murine and insect host models, the isolate was virulent. While information disseminated in the popular press would suggest this fungal contaminant poses little or no risk to consumers, our results show instead that it is capable of causing significant infections in animals