Mining the Pseudomonas genome Winsor, Geoffrey L; Brinkman, Fiona S L
Methods in molecular biology (Clifton, N.J.),
2014, Letnik:
1149
Journal Article
Pseudomonas species were targeted early for genomic studies since they were noted for their diverse metabolic capacity, ability to inhabit a wide range of environments and hosts, and include notable ...human and agriculturally relevant pathogens. As more genomes are sequenced, the power of genome-scale analyses are increasing and a wide range of analyses are now possible. The Pseudomonas Genome database has contributed to this effort by providing peer-reviewed, continually updated annotations of the Pseudomonas aeruginosa PAO1 reference strain genome plus integrated data and analyses of related Pseudomonas species. Analyses are now available via multiple resources to facilitate identification and characterization of drug targets, virulence factors, regulatory elements, genomic islands, genome rearrangements, orthologs, single nucleotide polymorphisms, and multiple other gene/protein-based analyses from gene expression to protein structure. We describe here how the Pseudomonas Genome Database and other bioinformatics resources can be leveraged to help Pseudomonas researchers "mine" Pseudomonas genomes, and associated genome-scale data, to facilitate new discovery.
The innate immune response is the first line of defence against invading pathogens and is regulated by complex signalling and transcriptional networks. Systems biology approaches promise to shed new ...light on the regulation of innate immunity through the analysis and modelling of these networks. A key initial step in this process is the contextual cataloguing of the components of this system and the molecular interactions that comprise these networks. InnateDB (http://www.innatedb.com) is a molecular interaction and pathway database developed to facilitate systems-level analyses of innate immunity.
Here, we describe the InnateDB curation project, which is manually annotating the human and mouse innate immunity interactome in rich contextual detail, and present our novel curation software system, which has been developed to ensure interactions are curated in a highly accurate and data-standards compliant manner. To date, over 13,000 interactions (protein, DNA and RNA) have been curated from the biomedical literature. Here, we present data, illustrating how InnateDB curation of the innate immunity interactome has greatly enhanced network and pathway annotation available for systems-level analysis and discuss the challenges that face such curation efforts. Significantly, we provide several lines of evidence that analysis of the innate immunity interactome has the potential to identify novel signalling, transcriptional and post-transcriptional regulators of innate immunity. Additionally, these analyses also provide insight into the cross-talk between innate immunity pathways and other biological processes, such as adaptive immunity, cancer and diabetes, and intriguingly, suggests links to other pathways, which as yet, have not been implicated in the innate immune response.
In summary, curation of the InnateDB interactome provides a wealth of information to enable systems-level analysis of innate immunity.
Toward the end of August 2000, the 6.3 Mbp whole genome sequence of Pseudomonas aeruginosa strain PAO1 was published. With 5570 open reading frames (ORFs), PAO1 had the largest microbial genome ...sequenced up to that point in time-including a large proportion of metabolic, transport and antimicrobial resistance genes supporting its ability to colonize diverse environments. A remarkable 9% of its ORFs were predicted to encode proteins with regulatory functions, providing new insight into bacterial network complexity as a function of network size. In this celebratory article, we fast forward 20 years, and examine how access to this resource has transformed our understanding of P. aeruginosa. What follows is more than a simple review or commentary; we have specifically asked some of the leaders in the field to provide personal reflections on how the PAO1 genome sequence, along with the Pseudomonas Community Annotation Project (PseudoCAP) and Pseudomonas Genome Database (pseudomonas.com), have contributed to the many exciting discoveries in this field. In addition to bringing us all up to date with the latest developments, we also ask our contributors to speculate on how the next 20 years of Pseudomonas research might pan out.
Many bacteria use the second messenger cyclic diguanylate (c-di-GMP) to control motility, biofilm production and virulence. Here, we identify a thermosensory diguanylate cyclase (TdcA) that modulates ...temperature-dependent motility, biofilm development and virulence in the opportunistic pathogen Pseudomonas aeruginosa. TdcA synthesizes c-di-GMP with catalytic rates that increase more than a hundred-fold over a ten-degree Celsius change. Analyses using protein chimeras indicate that heat-sensing is mediated by a thermosensitive Per-Arnt-SIM (PAS) domain. TdcA homologs are widespread in sequence databases, and a distantly related, heterologously expressed homolog from the Betaproteobacteria order Gallionellales also displayed thermosensitive diguanylate cyclase activity. We propose, therefore, that thermotransduction is a conserved function of c-di-GMP signaling networks, and that thermosensitive catalysis of a second messenger constitutes a mechanism for thermal sensing in bacteria.
1. Mating behaviour in Daphnia appears to rely on random contact between males and sexual females rather than diffusible pheromones. Males may be able to discriminate sexually receptive females from ...females in other developmental stages and increase their mating efficiency. Males may also use chemical signals to avoid mating with females from the same clone and avoid the severe inbreeding depression that has been documented for intraclonal mating. The present study used experiments to test for the avoidance of intraclonal mating and assess male mating efficiency in D. pulex.
2. Three clones were examined for the avoidance of intraclonal mating by providing males with an opportunity to mate with females of the same or two different clones. The proportion of intraclonal matings did not differ from the proportion of interclonal matings, suggesting that D. pulex males do not use kin discrimination to avoid mating with females from the same clone.
3. The proportion of mated females decreased with increasing numbers and density of sexual females when exposed to a single male. This observation suggests that a male spends more time pursuing and copulating with sexually receptive females than non‐receptive females and there is insufficient time to mate with all sexual females. The decrease in proportion of females mated could also be the result of sperm depletion in the male. Sperm depletion is unlikely to occur in nature because sexually receptive females are much rarer than in the experimental conditions.
The intestinal epithelial cell (IEC) represents the first cellular barrier to infection. Consistent with this sentinel role, IEC are known to produce a variety of chemokines in response to bacterial ...infection or proinflammatory cytokines. These chemokines act as potent leukocyte activators and chemoattractants in vivo. In this report, we begin to characterize the regulation of expression of the chemokine monocyte chemoattractant protein-1 (MCP-1) in the rat small intestinal IEC-18 line. Following stimulation with either interleukin-1beta (IL-1beta) or lipopolysaccharide (LPS), IEC-18 cells produced MCP-1, with IL-1 proving a more effective stimulus than LPS at both the mRNA and protein levels. Expression of MCP-1 due to either stimulus was inhibited by tyrosine kinase inhibitors, prompting us to investigate potential phosphotyrosine-dependent targets responsible for MCP-1 expression. We detected activation of p38, a member of the mitogen-activated protein kinase family, following either IL-1 or LPS treatment. Specific inhibition of this kinase using the compound SB203580 caused a destabilization of MCP-1 mRNA. These data point to a role for p38 in the regulation of MCP-1 mRNA expression by the IEC.
When the intestine becomes infected by pathogenic organisms, intestinal epithelial cells (IEC) respond with the production of chemokines, which then attract and activate specific subsets of ...leukocytes. During chronic inflammation, the panel of IEC chemokines produced likely represents the net effect of a plethora of mediators present in the milieu, including cytokines from activated T lymphocytes. To explore the influence of T lymphocyte cytokines, we treated IEC-18 cells with interferon-y (IFN-gamma) and interleukin-4 (IL-4) and measured the effect on production of the CC chemokines, monocyte chemoattractant protein-1 (MCP-1) and eotaxin, and the CXC chemokine, macrophage inflammatory protein-2 (MIP-2). Both IFN-gamma and IL-4 enhanced MCP-1 mRNA levels but with different kinetics. IFN-gamma stimulated a transient increase in MCP-1 mRNA levels, which peaked at 2 h, whereas IL-4-stimulated MCP-1 mRNA levels were markedly increased at 1 h and remained elevated at all time points studied. With each stimulus, the increase in MCP-1 mRNA levels was accompanied by a steady time-dependent increase in MCP-1 secretion. In addition, treatment with IFN-gamma or IL-4 enhanced IL-1beta-stimulated MCP-1 mRNA production and protein secretion. Eotaxin mRNA was detectable in unstimulated IEC-18 cells, and IL-4 but not IFN-gamma caused a rapid enhancement in levels, which remained elevated for 24 h after treatment. Finally, IL-1beta but not IFN-gamma or IL-4 enhanced MIP-2 mRNA levels. Knowledge gained from studying the outcome of T lymphocyte-derived stimuli will help understand the complex sequence of events during chronic intestinal inflammation.
Asexual organisms are thought to gain an advantage by avoiding the cost of producing males. In the cladoceran Daphnia pulex (Leydig), male production is determined by the environment and is ...independent of the origin of the asexual obligate parthenogens from the sexual cyclical parthenogens. If there is a cost to producing males, successful obligate parthenogens should have reduced or eliminated male production. Field and laboratory observations showed that obligate parthenogens have much-reduced male production compared to cyclical parthenogens. Although the reduction or elimination of males in the obligate parthenogens suggests that the cost of males is avoided, the coexistence of sexual and asexual forms of D. pulex may be partially explained by cyclical parthenogens compensating for the cost of males by having greater fecundity. In addition, the absence of a mating constraint for the obligate parthenogens may favour an increased allocation to asexual diapausing eggs earlier in the season compared to the cyclical parthenogens which require mating with males to produce sexual diapausing eggs. No difference in the production of diapausing eggs was observed, probably because males were abundant in populations of cyclical parthenogens and do not appear to limit the production of sexual diapausing eggs. D. pulex is a useful system for determining the ecological consequences of abandoning sexual reproduction and explaining the coexistence of sexual and asexual forms of a species.
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