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Despite enormous progress in understanding the fundamentals of bacterial gene regulation, our knowledge remains limited when compared with the number of bacterial genomes and regulatory systems to be discovered. Derived from a small number of initial studies, classic definitions for concepts of gene regulation have evolved as the number of characterized promoters has increased. Together with discoveries made using new technologies, this knowledge has led to revised generalizations and principles. In this Expert Recommendation, we suggest precise, updated definitions that support a logical, consistent conceptual framework of bacterial gene regulation, focusing on transcription initiation. The resulting concepts can be formalized by ontologies for computational modelling, laying the foundation for improved bioinformatics tools, knowledge-based resources and scientific communication. Thus, this work will help researchers construct better predictive models, with different formalisms, that will be useful in engineering, synthetic biology, microbiology and genetics.

BACKGROUND:Our understanding of the regulation of gene expression has benefited from the availability of high-throughput technologies that interrogate the whole genome for the binding of specific transcription factors and gene expression profiles. In the case of widely used model organisms, such as Escherichia coli K-12, the new knowledge gained from these approaches needs to be integrated with the legacy of accumulated knowledge from genetic and molecular biology experiments conducted in the pre-genomic era in order to attain the deepest level of understanding possible based on the available data. RESULTS:In this paper, we describe an expansion of RegulonDB, the database containing the rich legacy of decades of classic molecular biology experiments supporting what we know about gene regulation and operon organization in E. coli K-12, to include the genome-wide dataset collections from 32 ChIP and 19 gSELEX publications, in addition to around 60 genome-wide expression profiles relevant to the functional significance of these datasets and used in their curation. Three essential features for the integration of this information coming from different methodological approaches are: first, a controlled vocabulary within an ontology for precisely defining growth conditions; second, the criteria to separate elements with enough evidence to consider them involved in gene regulation from isolated transcription factor binding sites without such support; and third, an expanded computational model supporting this knowledge. Altogether, this constitutes the basis for adequately gathering and enabling the comparisons and integration needed to manage and access such wealth of knowledge. CONCLUSIONS:This version 10.0 of RegulonDB is a first step toward what should become the unifying access point for current and future knowledge on gene regulation in E. coli K-12. Furthermore, this model platform and associated methodologies and criteria can be emulated for gathering knowledge on other microbial organisms.

EcoCyc (http://EcoCyc.org) is a model organism database built on the genome sequence of Escherichia coli K-12 MG1655. Expert manual curation of the functions of individual E. coli gene products in EcoCyc has been based on information found in the experimental literature for E. coli K-12-derived strains. Updates to EcoCyc content continue to improve the comprehensive picture of E. coli biology. The utility of EcoCyc is enhanced by new tools available on the EcoCyc web site, and the development of EcoCyc as a teaching tool is increasing the impact of the knowledge collected in EcoCyc.

EcoCyc (http://EcoCyc.org) is a comprehensive model organism database for Escherichia coli K-12 MG1655. From the scientific literature, EcoCyc captures the functions of individual E. coli gene products; their regulation at the transcriptional, post-transcriptional and protein level; and their organization into operons, complexes and pathways. EcoCyc users can search and browse the information in multiple ways. Recent improvements to the EcoCyc Web interface include combined gene/protein pages and a Regulation Summary Diagram displaying a graphical overview of all known regulatory inputs to gene expression and protein activity. The graphical representation of signal transduction pathways has been updated, and the cellular and regulatory overviews were enhanced with new functionality. A specialized undergraduate teaching resource using EcoCyc is being developed.

EcoCyc (http://EcoCyc.org) provides a comprehensive encyclopedia of Escherichia coli biology. EcoCyc integrates information about the genome, genes and gene products; the metabolic network; and the regulatory network of E. coli. Recent EcoCyc developments include a new initiative to represent and curate all types of E. coli regulatory processes such as attenuation and regulation by small RNAs. EcoCyc has started to curate Gene Ontology (GO) terms for E. coli and has made a dataset of E. coli GO terms available through the GO Web site. The curation and visualization of electron transfer processes has been significantly improved. Other software and Web site enhancements include the addition of tracks to the EcoCyc genome browser, in particular a type of track designed for the display of ChIP-chip datasets, and the development of a comparative genome browser. A new Genome Omics Viewer enables users to paint omics datasets onto the full E. coli genome for analysis. A new advanced query page guides users in interactively constructing complex database queries against EcoCyc. A Macintosh version of EcoCyc is now available. A series of Webinars is available to instruct users in the use of EcoCyc.

The selective mechanisms operating in regulatory regions of bacterial genomes are poorly understood. In Escherichia coli, we have found that regulatory regions contain high densities of overlapping and probably competing promoter-like signals, in contrast to coding regions and regions located between convergently-transcribed genes. Functional promoter sites identified experimentally are often found in the subregions of highest density of signals, even when individual sites with higher binding affinity for RNA polymerase exist elsewhere within the region [Huerta & Collado-Vides 2003]. In order to explore whether this trend discovered in E. coli promoter regions is common in other bacterial species, we conducted similar analyses for a representative set containing 40 additional genomes belonging to different genera across all major bacterial phyla. This comparison is validated by the fact that RNAPs are evolutionarily conserved across all bacteria.

Bacteria of the genus Rhizobium and related genera establish nitrogen-fixing symbioses with the roots of leguminous plants. The genetic elements that participate in the symbiotic process are usually compartmentalized in the genome, either as independent replicons (symbiotic plasmids) or as symbiotic regions or islands in the chromosome. The complete nucleotide sequence of the symbiotic plasmid of Rhizobium etli model strain CFN42, symbiont of the common bean plant, has been reported. To better understand the basis of DNA sequence diversification of this symbiotic compartment, we analyzed the distribution of single-nucleotide polymorphisms in homologous regions from different Rhizobium etli strains. The distribution of polymorphisms is highly asymmetric in each of the different strains, alternating regions containing very few changes with regions harboring an elevated number of substitutions. The regions showing high polymorphism do not correspond with discrete genetic elements and are not the same in the different strains, indicating that they are not hypervariable regions of functional genes. Most interesting, some highly polymorphic regions share exactly the same nucleotide substitutions in more than one strain. Furthermore, in different regions of the symbiotic compartment, different sets of strains share the same substitutions. The data indicate that the majority of nucleotide substitutions are spread in the population by recombination and that the contribution of new mutations to polymorphism is relatively low. We propose that the horizontal transfer of homologous DNA segments among closely related organisms is a major source of genomic diversification.

Research has been conducted on the bacteriophage lambda promoters. The characterization of the interference dependence on the interpromoter distance is presented.