Unusual Regulation of a Leaderless Operon Involved in the Catabolism of Dimethylsulfoniopropionate in Rhodobacter sphaeroides

Rhodobacter sphaeroides strain 2.4.1 is a widely studied bacterium that has recently been shown to cleave the abundant marine anti-stress molecule dimethylsulfoniopropionate (DMSP) into acrylate plus gaseous dimethyl sulfide. It does so by using a lyase encoded by dddL, the promoter-distal gene of a three-gene operon, acuR-acuI-dddL. Transcription of the operon was enhanced when cells were pre-grown with the substrate DMSP, but this induction is indirect, and requires the conversion of DMSP to the product acrylate, the bona fide co-inducer. This regulation is mediated by the product of the promoter-proximal gene acuR, a transcriptional regulator in the TetR family. AcuR represses the operon in the absence of acrylate, but this is relieved by the presence of the co-inducer. Another unusual regulatory feature is that the acuR-acuI-dddL mRNA transcript is leaderless, such that acuR lacks a Shine-Dalgarno ribosomal binding site and 5′-UTR, and is translated at a lower level compared to the downstream genes. This regulatory unit may be quite widespread in bacteria, since several other taxonomically diverse lineages have adjacent acuR-like and acuI-like genes; these operons also have no 5′ leader sequences or ribosomal binding sites and their predicted cis-acting regulatory sequences resemble those of R. sphaeroides acuR-acuI-dddL

Tryptophan Scanning Analysis of the Membrane Domain of CTR-Copper Transporters

Membrane proteins of the CTR family mediate cellular copper uptake in all eukaryotic cells and have been shown to participate in uptake of platinum-based anticancer drugs. Despite their importance for life and the clinical treatment of malignancies, directed biochemical studies of CTR proteins have been difficult because high-resolution structural information is missing. Building on our recent 7Å structure of the human copper transporter hCTR1, we present the results of an extensive tryptophan-scanning analysis of hCTR1 and its distant relative, yeast CTR3. The comparative analysis supports our previous assignment of the transmembrane helices and shows that most functionally and structurally important residues are clustered around the threefold axis of CTR trimers or engage in helix packing interactions. The scan also identified residues that may play roles in interactions between CTR trimers and suggested that the first transmembrane helix serves as an adaptor that allows evolutionarily diverse CTRs to adopt the same overall structure. Together with previous biochemical and biophysical data, the results of the tryptophan scan are consistent with a mechanistic model in which copper transport occurs along the center of the trimer

Visualizing Interactions along the Escherichia coli Twin-Arginine Translocation Pathway Using Protein Fragment Complementation

The twin-arginine translocation (Tat) pathway is well known for its ability to export fully folded substrate proteins out of the cytoplasm of Gram-negative and Gram-positive bacteria. Studies of this mechanism in Escherichia coli have identified numerous transient protein-protein interactions that guide export-competent proteins through the Tat pathway. To visualize these interactions, we have adapted bimolecular fluorescence complementation (BiFC) to detect protein-protein interactions along the Tat pathway of living cells. Fragments of the yellow fluorescent protein (YFP) were fused to soluble and transmembrane factors that participate in the translocation process including Tat substrates, Tat-specific proofreading chaperones and the integral membrane proteins TatABC that form the translocase. Fluorescence analysis of these YFP chimeras revealed a wide range of interactions such as the one between the Tat substrate dimethyl sulfoxide reductase (DmsA) and its dedicated proofreading chaperone DmsD. In addition, BiFC analysis illuminated homo- and hetero-oligomeric complexes of the TatA, TatB and TatC integral membrane proteins that were consistent with the current model of translocase assembly. In the case of TatBC assemblies, we provide the first evidence that these complexes are co-localized at the cell poles. Finally, we used this BiFC approach to capture interactions between the putative Tat receptor complex formed by TatBC and the DmsA substrate or its dedicated chaperone DmsD. Our results demonstrate that BiFC is a powerful approach for studying cytoplasmic and inner membrane interactions underlying bacterial secretory pathways

Time-Resolved Transcriptome Analysis of Bacillus subtilis Responding to Valine, Glutamate, and Glutamine

Microorganisms can restructure their transcriptional output to adapt to environmental conditions by sensing endogenous metabolite pools. In this paper, an Agilent customized microarray representing 4,106 genes was used to study temporal transcript profiles of Bacillus subtilis in response to valine, glutamate and glutamine pulses over 24 h. A total of 673, 835, and 1135 amino-acid-regulated genes were identified having significantly changed expression at one or more time points in response to valine, glutamate, and glutamine, respectively, including genes involved in cell wall, cellular import, metabolism of amino-acids and nucleotides, transcriptional regulation, flagellar motility, chemotaxis, phage proteins, sporulation, and many genes of unknown function. Different amino acid treatments were compared in terms of both the global temporal profiles and the 5-minute quick regulations, and between-experiment differential genes were identified. The highlighted genes were analyzed based on diverse sources of gene functions using a variety of computational tools, including T-profiler analysis, and hierarchical clustering. The results revealed the common and distinct modes of action of these three amino acids, and should help to elucidate the specific signaling mechanism of each amino acid as an effector

Neurogenic mechanisms in bladder and bowel ageing

The prevalence of both urinary and faecal incontinence, and also chronic constipation, increases with ageing and these conditions have a major impact on the quality of life of the elderly. Management of bladder and bowel dysfunction in the elderly is currently far from ideal and also carries a significant financial burden. Understanding how these changes occur is thus a major priority in biogerontology. The functions of the bladder and terminal bowel are regulated by complex neuronal networks. In particular neurons of the spinal cord and peripheral ganglia play a key role in regulating micturition and defaecation reflexes as well as promoting continence. In this review we discuss the evidence for ageing-induced neuronal dysfunction that might predispose to neurogenic forms of incontinence in the elderly

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency–Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe