Metabolic analysis of the soil microbe Dechloromonas aromatica str. RCB: indications of a surprisingly complex life-style and cryptic anaerobic pathways for aromatic degradation

<p>Abstract</p> <p>Background</p> <p>Initial interest in <it>Dechloromonas aromatica </it>strain RCB arose from its ability to anaerobically degrade benzene. It is also able to reduce perchlorate and oxidize chlorobenzoate, toluene, and xylene, creating interest in using this organism for bioremediation. Little physiological data has been published for this microbe. It is considered to be a free-living organism.</p> <p>Results</p> <p>The <it>a priori </it>prediction that the <it>D. aromatica </it>genome would contain previously characterized "central" enzymes to support anaerobic aromatic degradation of benzene proved to be false, suggesting the presence of novel anaerobic aromatic degradation pathways in this species. These missing pathways include the benzylsuccinate synthase (<it>bss</it>ABC) genes (responsible for fumarate addition to toluene) and the central benzoyl-CoA pathway for monoaromatics. In depth analyses using existing TIGRfam, COG, and InterPro models, and the creation of <it>de novo </it>HMM models, indicate a highly complex lifestyle with a large number of environmental sensors and signaling pathways, including a relatively large number of GGDEF domain signal receptors and multiple quorum sensors. A number of proteins indicate interactions with an as yet unknown host, as indicated by the presence of predicted cell host remodeling enzymes, effector enzymes, hemolysin-like proteins, adhesins, NO reductase, and both type III and type VI secretory complexes. Evidence of biofilm formation including a proposed exopolysaccharide complex and exosortase (epsH) are also present. Annotation described in this paper also reveals evidence for several metabolic pathways that have yet to be observed experimentally, including a sulphur oxidation (<it>sox</it>FCDYZAXB) gene cluster, Calvin cycle enzymes, and proteins involved in nitrogen fixation in other species (including RubisCo, ribulose-phosphate 3-epimerase, and nif gene families, respectively).</p> <p>Conclusion</p> <p>Analysis of the <it>D. aromatica </it>genome indicates there is much to be learned regarding the metabolic capabilities, and life-style, for this microbial species. Examples of recent gene duplication events in signaling as well as dioxygenase clusters are present, indicating selective gene family expansion as a relatively recent event in <it>D. aromatica</it>'s evolutionary history. Gene families that constitute metabolic cycles presumed to create <it>D. aromatica'</it>s environmental 'foot-print' indicate a high level of diversification between its predicted capabilities and those of its close relatives, <it>A. aromaticum </it>str EbN1 and <it>Azoarcus </it>BH72.</p

Polarization of PAR Proteins by Advective Triggering of a Pattern-Forming System

In the Caenorhabditis elegans zygote, a conserved network of partitioning-defective (PAR) polarity proteins segregates into an anterior and a posterior domain, facilitated by flows of the cortical actomyosin meshwork. The physical mechanisms by which stable asymmetric PAR distributions arise from transient cortical flows remain unclear. We present evidence that PAR polarity arises from coupling of advective transport by the flowing cell cortex to a multistable PAR reaction-diffusion system. By inducing transient PAR segregation, advection serves as a mechanical trigger for the formation of a PAR pattern within an otherwise stably unpolarized system. We suggest that passive advective transport in an active and flowing material may be a general mechanism for mechanochemical pattern formation in developmental systems

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Parameter-space topology of models for cell polarity

Reaction-diffusion systems have been widely successful in the theoretical description of biological patterning phenomena, giving rise to numerous models based on differing mechanisms, mathematical implementations and parameter choices. However, even for models with common design features, the diversity of mathematical realizations may hinder the identification of common behavior. Here, we analyze three different reaction-diffusion models for cell polarity that feature conservation of mass, rapid cytoplasmic diffusion and bistability via a cusp bifurcation of uniform states. In all three models, the nonuniform polar states are front solutions, and growth of domains ceases through stalling of a propagating front. For these three models we find a characteristic parameter space topology, comprising a region of linear instability that loops around the cusp point and that is enclosed by a 'comet-shaped' region of nonuniform domain states. We propose a minimal model based on the cusp bifurcation normal form that includes essential characteristics of all cell polarity models considered. For this minimal model, we provide a complete analytical description of the parameter space topology, and find that the instability loop appears as a generic property of the cusp bifurcation. This topological analysis provides a unifying understanding of earlier mathematically distinct models and is suitable to classify future models. © 2014 IOP Publishing Ltd and Deutsche Physikalische Gesellschaft.SWG acknowledges funding from the European Research Council/ERC Grant agreement no 281903.Peer Reviewe

Final Report: Structural studies of archatelthermophilic adenylate kinase, September 15, 1996 - September 14, 1998

Through this DOE sponsored program Konisky has studied the evolution and molecular biology of microbes that live in extreme environments. The emphasis of this work has been the determination of the structural features of thermophilic enzymes that allow them to function optimally at near 100%. The laboratory has focused on a comparative study of adenylate kinase (ADK), an enzyme that functions to interconvert adenine nucleotides. Because of the close phylogenetic relatedness of members of the methanococci, differences in the structure of their ADKs will be dominated by structural features that reflect contributions to their optimal temperature for activity, rather than differences due to phylogenetic divergence. The authors have cloned, sequenced and modeled the secondary structure for several methanococcal ADKs. using molecular modeling threading approaches that are based on the solved structure for the porcine ADK, they have also proposed a general low resolution three dimensional structure for each of the methanococcal enzymes. These analyzes have allowed them to propose structural features that confer hyperthermoactivity to those enzymes functioning in the hyperthermophilic members of the Methanococci. Using protein engineering methodologies, they have tested their hypotheses by examining the effects of selective structural changes on thermoactivity. Despite possessing between 68--83% sequence identity, the methanococcal AKs had significantly different stability against thermal denaturation, with melting points ranging from 69--103 C. The construction of several chimerical AKs by linking regions of the MVO and MJA AKs demonstrated the importance of cooperative interactions between amino- and carboxyl-terminal regions in influencing thermostability. Addition of MJA terminal fragments to the MVO AK increased thermal stability approximately 20 C while maintaining 88% of the mesophilic sequence. Further analysis using structural models suggested that hydrophobic interactions are largely responsible for determining the thermostability of the methanococcal AKs. Construction of chimerical enzyme also demonstrated a distinct separation between thermostability and enzymatic temperature optima, suggesting that overall protein flexibility and stability are not dependently linked. Sequence comparisons and model buildings of highly related archaeal adenylate kinases has allowed the prediction of interactions responsible for the large temperature variation in temperatures for of optimal catalytic activity and temperature stability. The tertiary structure for these ADK have been predicted by using homology modeling to further investigate the potential of specific interactions on thermal stability and activity

GAViT: Genome Assembly Visualization Tool for Short Read Data

It is a challenging job for genome analysts to accurately debug, troubleshoot, and validate genome assembly results. Genome analysts rely on visualization tools to help validate and troubleshoot assembly results, including such problems as mis-assemblies, low-quality regions, and repeats. Short read data adds further complexity and makes it extremely challenging for the visualization tools to scale and to view all needed assembly information. As a result, there is a need for a visualization tool that can scale to display assembly data from the new sequencing technologies. We present Genome Assembly Visualization Tool (GAViT), a highly scalable and interactive assembly visualization tool developed at the DOE Joint Genome Institute (JGI)

In-silico Methods for Troubleshooting Genomic Shotgun Data

End-sequencing of shotgun libraries of small genomic inserts is today the most popular approach to Whole Genome Sequencing (WGS). Irregularities in WGS datasets present assembly problems that are expensive and time-consuming to solve, with cloning bias, contamination and long repeats posing the biggest challenges. Shotgun assembly data exhibit well recognizable patterns that follow certain statistical models, and deviations from these models usually stem from flaws and anomalies in the input data, which in turn reflect problems in the cloning protocol, chemistries, or the DNA being sequenced. We developed several statistical and bioinformatic methods for detecting cloning bias, DNA contamination and high repeat content at early stages of the WGS project. These methods are based on analyses of i) depth of coverage distributions, ii) dynamics of iterarative assemblies and iii) GC profiles of real and simulated shotgun datasets. We identify and describe relationships between read coverage and the Poisson function and demonstrate ways to routinely identify cloning bias and contamination through these relationships. Identifying abnormal patterns in the dataset s GC profile at various levels (s.a. genome, library, plate) provided a convenient method for catching suspected contamination. Routine automated application is also discussed

Production workflow tracking and QC analysis

The Joint Genome Institute Production Genomics Facility (PGF) has produced over 2.75 billion bases of draft paired-end sequencing since January 1, 2001. Our sequencing methodologies incorporate two types of DNA template generation: inoculation/SPRI purification and Rolling Circle Amplification (RCA). In order to manage the flow of samples through these processes, a robust database tracking system was developed using ORACLE. Data input and reporting for the workflow has been produced using a combination of commercial database development software and in-house programs. These include ORACLE s WebDB and CGI Perl programming. By leveraging the rapid report and form development cycle using WebDB and augmenting this with the flexibility of in-house programming, we have efficiently deployed a critical laboratory information management system for our data tracking