Mapping the DnaA-binding landscape reveals a preference for binding pairs of closely spaced DNA sites.

DnaA is a widely conserved DNA-binding protein that is essential for the initiation of DNA replication in many bacterial species, including . Cooperative binding of ATP-bound DnaA to multiple 9mer sites ('DnaA boxes') at the origin of replication results in local unwinding of the DNA and recruitment of the replication machinery. DnaA also functions as a transcription regulator by binding to DNA sites upstream of target genes.

Predictive Biophysical Neural Network Modeling of a Compendium of Transcription Factor DNA Binding Profiles for .

The DNA binding of most Transcription Factors (TFs) has not been comprehensively mapped, and few have models that can quantitatively predict binding affinity. We report the global mapping of DNA binding for 139 TFs using ChIP-Seq. We used these data to train BoltzNet, a novel neural network that predicts TF binding energy from DNA sequence.

Direct and indirect control of Rho-dependent transcription termination by the riboswitch.

Bacterial riboswitches are molecular structures that play a crucial role in controlling gene expression to maintain cellular balance. The riboswitch has been previously shown to regulate gene expression through translation initiation and mRNA decay. Recent research suggests that gene expression is also influenced by Rho-dependent transcription termination.

Promoter selectivity of the RhlR quorum-sensing transcription factor receptor in Pseudomonas aeruginosa is coordinated by distinct and overlapping dependencies on C4-homoserine lactone and PqsE.

Quorum sensing is a mechanism of bacterial cell-cell communication that relies on the production and detection of small molecule autoinducers, which facilitate the synchronous expression of genes involved in group behaviors, such as virulence factor production and biofilm formation. The Pseudomonas aeruginosa quorum sensing network consists of multiple interconnected transcriptional regulators, with the transcription factor, RhlR, acting as one of the main drivers of quorum sensing behaviors. RhlR is a LuxR-type transcription factor that regulates its target genes when bound to its cognate autoinducer, C4-homoserine lactone, which is synthesized by RhlI.

Mapping direct and indirect MarA/SoxS/Rob/RamA regulons in Typhimurium reveals repression of and biofilm formation.

The closely related transcription factors MarA, SoxS, Rob and RamA control overlapping stress responses in many enteric bacteria. Furthermore, constitutive expression of such regulators is linked to clinical antibiotic resistance. In this work we have mapped the binding of MarA, SoxS, Rob and RamA across the Typhimurium genome.

Genome-Wide Mapping of the Escherichia coli PhoB Regulon Reveals Many Transcriptionally Inert, Intragenic Binding Sites.

Genome-scale analyses have revealed many transcription factor binding sites within, rather than upstream of, genes, raising questions as to the function of these binding sites. Here, we use complementary approaches to map the regulon of the Escherichia coli transcription factor PhoB, a response regulator that controls transcription of genes involved in phosphate homeostasis. Strikingly, the majority of PhoB binding sites are located within genes, but these intragenic sites are not associated with detectable transcription regulation and are not evolutionarily conserved.

The small mycobacterial ribosomal protein, bS22, modulates aminoglycoside accessibility to its 16S rRNA helix-44 binding site.

Treatment of tuberculosis continues to be challenging due to the widespread latent form of the disease and the emergence of antibiotic-resistant strains of the pathogen, . Bacterial ribosomes are a common and effective target for antibiotics. Several second line anti-tuberculosis drugs, e.

Genome-wide mapping of the PhoB regulon reveals many transcriptionally inert, intragenic binding sites.

Unlabelled: Genome-scale analyses have revealed many transcription factor binding sites within, rather than upstream of genes, raising questions as to the function of these binding sites. Here, we use complementary approaches to map the regulon of the transcription factor PhoB, a response regulator that controls transcription of genes involved in phosphate homeostasis. Strikingly, the majority of PhoB binding sites are located within genes, but these intragenic sites are not associated with detectable transcription regulation and are not evolutionarily conserved.

Transcription-Translation Coupling in Bacteria.

In bacteria, transcription and translation take place in the same cellular compartment. Therefore, a messenger RNA can be translated as it is being transcribed, a process known as transcription-translation coupling. This process was already recognized at the dawn of molecular biology, yet the interplay between the two key players, the RNA polymerase and ribosome, remains elusive.

RegulonDB 11.0: Comprehensive high-throughput datasets on transcriptional regulation in K-12.

Genomics has set the basis for a variety of methodologies that produce high-throughput datasets identifying the different players that define gene regulation, particularly regulation of transcription initiation and operon organization. These datasets are available in public repositories, such as the Gene Expression Omnibus, or ArrayExpress. However, accessing and navigating such a wealth of data is not straightforward.

Pervasive translation in .

Most bacterial ORFs are identified by automated prediction algorithms. However, these algorithms often fail to identify ORFs lacking canonical features such as a length of >50 codons or the presence of an upstream Shine-Dalgarno sequence. Here, we use ribosome profiling approaches to identify actively translated ORFs in .

Spatiotemporal localization of proteins in mycobacteria.

Although prokaryotic organisms lack traditional organelles, they must still organize cellular structures in space and time, challenges that different species solve differently. To systematically define the subcellular architecture of mycobacteria, we perform high-throughput imaging of a library of fluorescently tagged proteins expressed in Mycobacterium smegmatis and develop a customized computational pipeline, MOMIA and GEMATRIA, to analyze these data. Our results establish a spatial organization network of over 700 conserved mycobacterial proteins and reveal a coherent localization pattern for many proteins of known function, including those in translation, energy metabolism, cell growth and division, as well as proteins of unknown function.

A Practical Guide to Small Protein Discovery and Characterization Using Mass Spectrometry.

Small proteins of up to ∼50 amino acids are an abundant class of biomolecules across all domains of life. Yet due to the challenges inherent in their size, they are often missed in genome annotations, and are difficult to identify and characterize using standard experimental approaches. Consequently, we still know few small proteins even in well-studied prokaryotic model organisms.

Fluorescence Imaging-Based Discovery of Membrane Domain-Associated Proteins in .

Mycobacteria spatially organize their plasma membrane, and many enzymes involved in envelope biosynthesis associate with a membrane compartment termed the intracellular membrane domain (IMD). The IMD is concentrated in the polar regions of growing cells and becomes less polarized under nongrowing conditions. Because mycobacteria elongate from the poles, the observed polar localization of the IMD during growth likely supports the localized biosynthesis of envelope components.

Dissecting Locus Regulation in Yersinia pestis.

The pH 6 antigen (PsaA) of Yersinia pestis is a virulence factor that is expressed in response to high temperature (37°C) and low pH (6.0). Previous studies have implicated the PsaE and PsaF regulators in the temperature- and pH-dependent regulation of .

Widespread divergent transcription from bacterial and archaeal promoters is a consequence of DNA-sequence symmetry.

Transcription initiates at promoters, DNA regions recognized by a DNA-dependent RNA polymerase. We previously identified horizontally acquired Escherichia coli promoters from which the direction of transcription was unclear. In the present study, we show that more than half of these promoters are bidirectional and drive divergent transcription.

Regulatory roles of 5' UTR and ORF-internal RNAs detected by 3' end mapping.

Many bacterial genes are regulated by RNA elements in their 5´ untranslated regions (UTRs). However, the full complement of these elements is not known even in the model bacterium . Using complementary RNA-sequencing approaches, we detected large numbers of 3´ ends in 5´ UTRs and open reading frames (ORFs), suggesting extensive regulation by premature transcription termination.

Regulatory Cross Talk between Motility and Interbacterial Communication in Salmonella enterica Serovar Typhimurium.

FliA is a broadly conserved σ factor that directs transcription of genes involved in flagellar motility. We previously identified FliA-transcribed genes in and serovar Typhimurium, and we showed that FliA transcribes many unstable, noncoding RNAs from intragenic promoters. Here, we show that FliA in Typhimurium also directs the transcription of large numbers of unstable, noncoding RNAs from intragenic promoters, and we identify two previously unreported FliA-transcribed protein-coding genes.

Transcription termination and antitermination of bacterial CRISPR arrays.

A hallmark of CRISPR-Cas immunity systems is the CRISPR array, a genomic locus consisting of short, repeated sequences ('repeats') interspersed with short, variable sequences ('spacers'). CRISPR arrays are transcribed and processed into individual CRISPR RNAs that each include a single spacer, and direct Cas proteins to complementary sequences in invading nucleic acid. Most bacterial CRISPR array transcripts are unusually long for untranslated RNA, suggesting the existence of mechanisms to prevent premature transcription termination by Rho, a conserved bacterial transcription termination factor that rapidly terminates untranslated RNA.

Redefining fundamental concepts of transcription initiation in bacteria.

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.

Transcriptional profiling of Vibrio cholerae O1 following exposure to human anti- lipopolysaccharide monoclonal antibodies.

Following an episode of cholera, a rapidly dehydrating, watery diarrhea caused by the Gram-negative bacterium, Vibrio cholerae O1, humans mount a robust anti-lipopolysaccharide (LPS) antibody response that is associated with immunity to subsequent re-infection. In neonatal mouse and rabbit models of cholera, passively administered anti-LPS polyclonal and monoclonal (MAb) antibodies reduce V. cholerae colonization of the intestinal epithelia by inhibiting bacterial motility and promoting vibrio agglutination.