NCBI GEO: archive for gene expression and epigenomics data sets: 23-year update.

The Gene Expression Omnibus (GEO) is an international public repository that archives gene expression and epigenomics data sets generated by next-generation sequencing and microarray technologies. Data are typically submitted to GEO by researchers in compliance with widespread journal and funder mandates to make generated data publicly accessible. The resource handles raw data files, processed data files and descriptive metadata for over 200 000 studies and 6.5 million samples, all of which are indexed, searchable and downloadable. Additionally, GEO offers web-based tools that facilitate analysis and visualization of differential gene expression. This article presents the current status and recent advancements in GEO, including the generation of consistently computed gene expression count matrices for thousands of RNA-seq studies, and new interactive graphical plots in GEO2R that help users identify differentially expressed genes and assess data set quality. The GEO repository is built and maintained by the National Center for Biotechnology Information (NCBI), a division of the National Library of Medicine (NLM), and is publicly accessible at https://www.ncbi.nlm.nih.gov/geo/.

NCBI GEO: archive for functional genomics data sets--update.

The Gene Expression Omnibus (GEO, http://www.ncbi.nlm.nih.gov/geo/) is an international public repository for high-throughput microarray and next-generation sequence functional genomic data sets submitted by the research community. The resource supports archiving of raw data, processed data and metadata which are indexed, cross-linked and searchable. All data are freely available for download in a variety of formats. GEO also provides several web-based tools and strategies to assist users to query, analyse and visualize data. This article reports current status and recent database developments, including the release of GEO2R, an R-based web application that helps users analyse GEO data.

NCBI GEO: archive for functional genomics data sets--10 years on.

A decade ago, the Gene Expression Omnibus (GEO) database was established at the National Center for Biotechnology Information (NCBI). The original objective of GEO was to serve as a public repository for high-throughput gene expression data generated mostly by microarray technology. However, the research community quickly applied microarrays to non-gene-expression studies, including examination of genome copy number variation and genome-wide profiling of DNA-binding proteins. Because the GEO database was designed with a flexible structure, it was possible to quickly adapt the repository to store these data types. More recently, as the microarray community switches to next-generation sequencing technologies, GEO has again adapted to host these data sets. Today, GEO stores over 20,000 microarray- and sequence-based functional genomics studies, and continues to handle the majority of direct high-throughput data submissions from the research community. Multiple mechanisms are provided to help users effectively search, browse, download and visualize the data at the level of individual genes or entire studies. This paper describes recent database enhancements, including new search and data representation tools, as well as a brief review of how the community uses GEO data. GEO is freely accessible at http://www.ncbi.nlm.nih.gov/geo/.

GeMInA, Genomic Metadata for Infectious Agents, a geospatial surveillance pathogen database.

The Gemina system (http://gemina.igs.umaryland.edu) identifies, standardizes and integrates the outbreak metadata for the breadth of NIAID category A-C viral and bacterial pathogens, thereby providing an investigative and surveillance tool describing the Who [Host], What [Disease, Symptom], When [Date], Where [Location] and How [Pathogen, Environmental Source, Reservoir, Transmission Method] for each pathogen. The Gemina database will provide a greater understanding of the interactions of viral and bacterial pathogens with their hosts and infectious diseases through in-depth literature text-mining, integrated outbreak metadata, outbreak surveillance tools, extensive ontology development, metadata curation and representative genomic sequence identification and standards development. The Gemina web interface provides metadata selection and retrieval of a pathogen's; Infection Systems (Pathogen, Host, Disease, Transmission Method and Anatomy) and Incidents (Location and Date) along with a hosts Age and Gender. The Gemina system provides an integrated investigative and geospatial surveillance system connecting pathogens, pathogen products and disease anchored on the taxonomic ID of the pathogen and host to identify the breadth of hosts and diseases known for these pathogens, to identify the extent of outbreak locations, and to identify unique genomic regions with the DNA Signature Insignia Detection Tool.

NCBI GEO: archive for high-throughput functional genomic data.

The Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI) is the largest public repository for high-throughput gene expression data. Additionally, GEO hosts other categories of high-throughput functional genomic data, including those that examine genome copy number variations, chromatin structure, methylation status and transcription factor binding. These data are generated by the research community using high-throughput technologies like microarrays and, more recently, next-generation sequencing. The database has a flexible infrastructure that can capture fully annotated raw and processed data, enabling compliance with major community-derived scientific reporting standards such as 'Minimum Information About a Microarray Experiment' (MIAME). In addition to serving as a centralized data storage hub, GEO offers many tools and features that allow users to effectively explore, analyze and download expression data from both gene-centric and experiment-centric perspectives. This article summarizes the GEO repository structure, content and operating procedures, as well as recently introduced data mining features. GEO is freely accessible at http://www.ncbi.nlm.nih.gov/geo/.

High-throughput phenotypic characterization of Pseudomonas aeruginosa membrane transport genes.

The deluge of data generated by genome sequencing has led to an increasing reliance on bioinformatic predictions, since the traditional experimental approach of characterizing gene function one at a time cannot possibly keep pace with the sequence-based discovery of novel genes. We have utilized Biolog phenotype MicroArrays to identify phenotypes of gene knockout mutants in the opportunistic pathogen and versatile soil bacterium Pseudomonas aeruginosa in a relatively high-throughput fashion. Seventy-eight P. aeruginosa mutants defective in predicted sugar and amino acid membrane transporter genes were screened and clear phenotypes were identified for 27 of these. In all cases, these phenotypes were confirmed by independent growth assays on minimal media. Using qRT-PCR, we demonstrate that the expression levels of 11 of these transporter genes were induced from 4- to 90-fold by their substrates identified via phenotype analysis. Overall, the experimental data showed the bioinformatic predictions to be largely correct in 22 out of 27 cases, and led to the identification of novel transporter genes and a potentially new histamine catabolic pathway. Thus, rapid phenotype identification assays are an invaluable tool for confirming and extending bioinformatic predictions.

Whole-genome microarray analyses of Synechococcus-Vibrio interactions.

Microbes live in diverse communities yet their physiologies are typically studied in axenic culture. To begin to address this dichotomy, whole-genome microarray analyses were used and revealed that several major metabolic pathways were affected in Synechococcus sp. WH8102, a model phototroph, when grown with Vibrio parahaemolyticus, a model heterotroph. In co-cultures with V. parahaemolyticus, although phosphate was not depleted, Synechococcus sp. WH8102 may have experienced phosphate stress since the expression of phosphate acquisition genes increased and alkaline phosphatase activity was higher than in monocultures. Expression of cell wall synthesis genes and the components of a zinc transporter were also upregulated. In contrast, a ferric uptake regulation (Fur) family gene was downregulated as were genes that encode proteins rich in iron or involved in detoxifying oxygen radicals. Nitrogen use may also have been affected in co-cultures as the gene expression changes share similarities with ammonia-grown Synechococcus. This study demonstrates the multiple impacts that interspecific microbial interactions can have on the physiology of a major primary producer and the importance of investigating microbial physiology from a community perspective.

Impact of DNA damaging agents on genome-wide transcriptional profiles in two marine Synechococcus species.

Marine microorganisms, particularly those residing in coastal areas, may come in contact with any number of chemicals of environmental or xenobiotic origin. The sensitivity and response of marine cyanobacteria to such chemicals is, at present, poorly understood. We have looked at the transcriptional response of well characterized Synechococcus open ocean (WH8102) and coastal (CC9311) isolates to two DNA damaging agents, mitomycin C and ethidium bromide, using whole-genome expression microarrays. The coastal strain showed differential regulation of a larger proportion of its genome following "shock" treatment with each agent. Many of the orthologous genes in these strains, including those encoding sensor kinases, showed different transcriptional responses, with the CC9311 genes more likely to show significant changes in both treatments. While the overall response of each strain was considerably different, there were distinct transcriptional responses common to both strains observed for each DNA damaging agent, linked to the mode of action of each chemical. In both CC9311 and WH8102 there was evidence of SOS response induction under mitomycin C treatment, with genes recA, lexA and umuC significantly upregulated in this experiment but not under ethidium bromide treatment. Conversely, ethidium bromide treatment tended to result in upregulation of the DNA-directed RNA polymerase genes, not observed following mitomycin C treatment. Interestingly, a large number of genes residing on putative genomic island regions of each genome also showed significant upregulation under one or both chemical treatments.

PtrA is required for coordinate regulation of gene expression during phosphate stress in a marine Synechococcus.

Previous microarray analyses have shown a key role for the two-component system PhoBR (SYNW0947, SYNW0948) in the regulation of P transport and metabolism in the marine cyanobacterium Synechococcus sp. WH8102. However, there is some evidence that another regulator, SYNW1019 (PtrA), probably under the control of PhoBR, is involved in the response to P depletion. PtrA is a member of the cAMP receptor protein transcriptional regulator family that shows homology to NtcA, the global nitrogen regulator in cyanobacteria. To define the role of this regulator, we constructed a mutant by insertional inactivation and compared the physiology of wild-type Synechcococcus sp. WH8102 with the ptrA mutant under P-replete and P-stress conditions. In response to P stress the ptrA mutant failed to upregulate phosphatase activity. Microarrays and quantitative RT-PCR indicate that a subset of the Pho regulon is controlled by PtrA, including two phosphatases, a predicted phytase and a gene of unknown function psip1 (SYNW0165), all of which are highly upregulated during P limitation. Electrophoretic mobility shift assays indicate binding of overexpressed PtrA to promoter sequences upstream of the induced genes. This work suggests a two-tiered response to P depletion in this strain, the first being PhoB-dependent induction of high-affinity PO(4) transporters, and the second the PtrA-dependent induction of phosphatases for scavenging organic P. The levels of numerous other transcripts are also directly or indirectly influenced by PtrA, including those involved in cell-surface modification, metal uptake, photosynthesis, stress responses and other metabolic processes, which may indicate a wider role for PtrA in cellular regulation in marine picocyanobacteria.

Microarray analysis of phosphate regulation in the marine cyanobacterium Synechococcus sp. WH8102.

Primary productivity of open ocean environments, such as those inhabited by marine picocyanobacteria, is often limited by low inorganic phosphate (P). To observe how these organisms cope with P starvation, we constructed a full genome microarray for Synechococcus sp. WH8102 and compared differences in gene expression under P-replete and P-limited growth conditions, including both early P stress, during extracellular alkaline phosphatase induction, and late P stress. A total of 36 genes showed significant upregulation (>log(2) fold) whereas 23 genes were highly downregulated at the early time point; however, these changes in expression were maintained during late P stress for only 5 of the upregulated genes. Knockout mutants were constructed for genes SYNW0947 and SYNW0948, comprising a two-component regulator hypothesized to have a key function in regulating P metabolism. A high degree of overlap in the sets of genes affected by P stress conditions and in the knockout mutants supports this hypothesis; however, there is some indication that other regulators may be involved in this response in Synechococcus sp. WH8102. Consistent with what has been observed in many other cyanobacteria, the Pho regulon of this strain is comprised largely of genes for alkaline phosphatases, P transport or P metabolism. Interestingly, however, the exact composition and arrangement of the Pho regulon appears highly variable in marine cyanobacteria.