Search

DNA transcription initiates after an RNA polymerase (RNAP) molecule binds to the promoter of a gene. In bacteria, the canonical picture is that RNAP comes from the cytoplasmic pool of freely diffusing RNAP molecules. Recent experiments suggest the possible existence of a separate pool of polymerases, competent for initiation, which freely slide on the DNA after having terminated one round of transcription. Promoter-dependent transcription reinitiation from this pool of posttermination RNAP may lead to coupled initiation at nearby operons, but it is unclear whether this can occur over the distance and timescales needed for it to function widely on a bacterial genome in vivo. Here, we mathematically model the hypothesized reinitiation mechanism as a diffusion-to-capture process and compute the distances over which significant interoperon coupling can occur and the time required. These quantities depend on molecular association and dissociation rate constants between DNA, RNAP, and the transcription initiation factor σ 70 ; we measure these rate constants using single-molecule experiments in vitro. Our combined theory/experimental results demonstrate that efficient coupling can occur at physiologically relevant σ 70 concentrations and on timescales appropriate for transcript synthesis. Coupling is efficient over terminator-promoter distances up to ∼1,000 bp, which includes the majority of terminator-promoter nearest neighbor pairs in the Escherichia coli genome. The results suggest a generalized mechanism that couples the transcription of nearby operons and breaks the paradigm that each binding of RNAP to DNA can produce at most one messenger RNA.

Free-living bacteria have regulatory systems that can quickly reprogram gene transcription in response to changes in the cellular environment. The RapA ATPase, a prokaryotic homolog of the eukaryotic Swi2/Snf2 chromatin remodeling complex, may facilitate such reprogramming, but the mechanisms by which it does so are unclear. We used multiwavelength single-molecule fluorescence microscopy in vitro to examine RapA function in the Escherichia coli transcription cycle. In our experiments, RapA at <5 nM concentration did not appear to alter transcription initiation, elongation, or intrinsic termination. Instead, we directly observed a single RapA molecule bind specifically to the kinetically stable post termination complex (PTC)-consisting of core RNA polymerase (RNAP)-bound sequence nonspecifically to double-stranded DNA-and efficiently remove RNAP from DNA within seconds in an ATP-hydrolysis-dependent reaction. Kinetic analysis elucidates the process through which RapA locates the PTC and the key mechanistic intermediates that bind and hydrolyze ATP. This study defines how RapA participates in the transcription cycle between termination and initiation and suggests that RapA helps set the balance between global RNAP recycling and local transcription reinitiation in proteobacterial genomes.

Bacillus cereus is a gram positive microorganism commonly involved in gastrointestinal infection but capable of causing severe infections and bacteremia. We describe here a case of bacteremia caused by B. cereus in a previously healthy young woman admitted to the intensive care unit following emergency surgery due to a penetrating abdominal stab wound and subsequent hepatic lesion. She developed fever during admission and cultures were taken. B. cereus was isolated in blood and hepatic fluid collection cultures. Treatment was adjusted according to the isolate, with good clinical results. It is important to highlight the pathogenic potential of this microorganism and not underestimate it as a contaminant when it is isolated from blood samples., (Copyright © 2019 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Biological systems are spatially organized. This microscopic heterogeneity has been shown to produce emergent complex behaviors such as bistability. Even though the connection between spatiality and dynamic response is essential to understand biological output, its robustness and extent has not been sufficiently explored. This work focuses on a previously described system which is composed of two monostable modules acting on different cellular compartments and sharing species through linear shuttling reactions. One of the two main purposes of this paper is to quantify the frequency of occurrence of bistability throughout the parameter space and to identify which parameters and in which value ranges control the emergence and the properties of bistability. We found that a very small fraction of the sampled parameter space produced a bistable response. Most importantly, shuttling parameters were among the most influential ones to control this property. The other goal of this paper is to simplify the same system as much as possible without losing compartment-induced bistability. This procedure provided a simplified model that still connects two monostable systems by a reduced set of linear shuttling reactions that circulates all the species around the two compartments. Bistable systems are one of the main building blocks of more complex behaviors such as oscillations, memory, and digitalization. Therefore, we expect that the proposed minimal system provides insight into how these behaviors can arise from compartmentalization.

Scedosporium is a fungus that has a worldwide distribution, and which can be found in soil and contaminated water. It can rarely affect bone tissue and can do it either by direct inoculation or through trauma. We present here a case of a 54- year- old male patient with a diagnosis of chronic bacterial osteomyelitis due to an aquatic accident and exposed fracture of tibia-fibula of both members, which was treated with broad-spectrum antibiotics for 120 days. Eight months after the onset of the disease, Scedosporium spp. was isolated from the collection of one of the affected member, which was treated with voriconazole in combination with terbinafine., (Copyright © 2019 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Human herpesviruses 6A and 6B (HHV-6A and HHV-6B) are human viruses capable of chromosomal integration. Approximately 1% of the human population carries one copy of HHV-6A/B integrated into every cell in their body, referred to as inherited chromosomally integrated human herpesvirus 6A/B (iciHHV-6A/B). Whether iciHHV-6A/B is transcriptionally active in vivo and how it shapes the immunological response are still unclear. In this study, we screened DNA sequencing (DNA-seq) and transcriptome sequencing (RNA-seq) data for 650 individuals available through the Genotype-Tissue Expression (GTEx) project and identified 2 iciHHV-6A- and 4 iciHHV-6B-positive candidates. When corresponding tissue-specific gene expression signatures were analyzed, low levels HHV-6A/B gene expression was found across multiple tissues, with the highest levels of gene expression in the brain (specifically for HHV-6A), testis, esophagus, and adrenal gland. U90 and U100 were the most highly expressed HHV-6 genes in both iciHHV-6A- and iciHHV-6B-positive individuals. To assess whether tissue-specific gene expression from iciHHV-6A/B influences the immune response, a cohort of 15,498 subjects was screened and 85 iciHHV-6A/B + subjects were identified. Plasma samples from iciHHV-6A/B + and age- and sex-matched controls were analyzed for antibodies to control antigens (cytomegalovirus [CMV], Epstein-Barr virus [EBV], and influenza virus [FLU]) or HHV-6A/B antigens. Our results indicate that iciHHV-6A/B + subjects have significantly more antibodies against the U90 gene product (IE1) than do non-iciHHV-6-positive individuals. Antibody responses against EBV and FLU antigens or HHV-6A/B gene products either not expressed or expressed at low levels, such as U47, U57, and U72, were identical between controls and iciHHV-6A/B + subjects. CMV-seropositive individuals with iciHHV-6A/B + have more antibodies against CMV pp150 than do CMV-seropositive controls. These results argue that spontaneous gene expression from integrated HHV-6A/B leads to an increase in antigenic burden that translates into a more robust HHV-6A/B-specific antibody response. IMPORTANCE HHV-6A and -6B are human herpesviruses that have the unique property of being able to integrate into the telomeric regions of human chromosomes. Approximately 1% of the world's population carries integrated HHV-6A/B genome in every cell of their body. Whether viral genes are transcriptionally active in these individuals is unclear. By taking advantage of a unique tissue-specific gene expression data set, we showed that the majority of tissues from iciHHV-6 individuals do not show HHV-6 gene expression. Brain and testes showed the highest tissue-specific expression of HHV-6 genes in two separate data sets. Two HHV-6 genes, U90 (immediate early 1 protein) and U100 (glycoproteins Q1 and Q2), were found to be selectively and consistently expressed across several human tissues. Expression of U90 translates into an increase in antigen-specific antibody response in iciHHV-6A/B + subjects relative to controls. Future studies will be needed to determine the mechanism of gene expression, the effects of these genes on human gene transcription networks, and the pathophysiological impact of having increased viral protein expression in tissue in conjunction with increased antigen-specific antibody production., (Copyright © 2019 Peddu et al.)

Background: CUT&RUN is an efficient epigenome profiling method that identifies sites of DNA binding protein enrichment genome-wide with high signal to noise and low sequencing requirements. Currently, the analysis of CUT&RUN data is complicated by its exceptionally low background, which renders programs designed for analysis of ChIP-seq data vulnerable to oversensitivity in identifying sites of protein binding., Results: Here we introduce Sparse Enrichment Analysis for CUT&RUN (SEACR), an analysis strategy that uses the global distribution of background signal to calibrate a simple threshold for peak calling. SEACR discriminates between true and false-positive peaks with near-perfect specificity from "gold standard" CUT&RUN datasets and efficiently identifies enriched regions for several different protein targets. We also introduce a web server ( http://seacr.fredhutch.org ) for plug-and-play analysis with SEACR that facilitates maximum accessibility across users of all skill levels., Conclusions: SEACR is a highly selective peak caller that definitively validates the accuracy of CUT&RUN for datasets with known true negatives. Its ease of use and performance in comparison with existing peak calling strategies make it an ideal choice for analyzing CUT&RUN data.

Bioconductor is an open-source, open-development software project for the analysis and comprehension of high-throughput data in genomics and molecular biology. The project aims to enable interdisciplinary research, collaboration and rapid development of scientific software. Based on the statistical programming language R, Bioconductor comprises 934 interoperable packages contributed by a large, diverse community of scientists. Packages cover a range of bioinformatic and statistical applications. They undergo formal initial review and continuous automated testing. We present an overview for prospective users and contributors.

Assembly of genes into operons is generally viewed as an important process during the continual adaptation of microbes to changing environmental challenges. However, the genome reorganization events that drive this process are also the roots of instability for existing operons. We have determined that there exists a statistically significant trend that correlates the proportion of genes encoded in operons in archaea to their phylogenetic lineage. We have further characterized how microbes deal with operon instability by mapping and comparing transcriptome architectures of four phylogenetically diverse extremophiles that span the range of operon stabilities observed across archaeal lineages: a photoheterotrophic halophile (Halobacterium salinarum NRC-1), a hydrogenotrophic methanogen (Methanococcus maripaludis S2), an acidophilic and aerobic thermophile (Sulfolobus solfataricus P2), and an anaerobic hyperthermophile (Pyrococcus furiosus DSM 3638). We demonstrate how the evolution of transcriptional elements (promoters and terminators) generates new operons, restores the coordinated regulation of translocated, inverted, and newly acquired genes, and introduces completely novel regulation for even some of the most conserved operonic genes such as those encoding subunits of the ribosome. The inverse correlation (r=-0.92) between the proportion of operons with such internally located transcriptional elements and the fraction of conserved operons in each of the four archaea reveals an unprecedented view into varying stages of operon evolution. Importantly, our integrated analysis has revealed that organisms adapted to higher growth temperatures have lower tolerance for genome reorganization events that disrupt operon structures.

Background: High-density tiling arrays and new sequencing technologies are generating rapidly increasing volumes of transcriptome and protein-DNA interaction data. Visualization and exploration of this data is critical to understanding the regulatory logic encoded in the genome by which the cell dynamically affects its physiology and interacts with its environment., Results: The Gaggle Genome Browser is a cross-platform desktop program for interactively visualizing high-throughput data in the context of the genome. Important features include dynamic panning and zooming, keyword search and open interoperability through the Gaggle framework. Users may bookmark locations on the genome with descriptive annotations and share these bookmarks with other users. The program handles large sets of user-generated data using an in-process database and leverages the facilities of SQL and the R environment for importing and manipulating data.A key aspect of the Gaggle Genome Browser is interoperability. By connecting to the Gaggle framework, the genome browser joins a suite of interconnected bioinformatics tools for analysis and visualization with connectivity to major public repositories of sequences, interactions and pathways. To this flexible environment for exploring and combining data, the Gaggle Genome Browser adds the ability to visualize diverse types of data in relation to its coordinates on the genome., Conclusions: Genomic coordinates function as a common key by which disparate biological data types can be related to one another. In the Gaggle Genome Browser, heterogeneous data are joined by their location on the genome to create information-rich visualizations yielding insight into genome organization, transcription and its regulation and, ultimately, a better understanding of the mechanisms that enable the cell to dynamically respond to its environment.

Gaggle Tool Creator (GTC) is a web application which provides access to public annotation, interaction, orthology, and genomic data for hundreds of organisms, and enables instant analysis of the data using many popular web-based and desktop applications.

High-throughput studies of biological systems are rapidly accumulating a wealth of 'omics'-scale data. Visualization is a key aspect of both the analysis and understanding of these data, and users now have many visualization methods and tools to choose from. The challenge is to create clear, meaningful and integrated visualizations that give biological insight, without being overwhelmed by the intrinsic complexity of the data. In this review, we discuss how visualization tools are being used to help interpret protein interaction, gene expression and metabolic profile data, and we highlight emerging new directions.