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Molecular Microbiology

Rahul Shrivastava, Xiang'Er Jiang, Shu-Sin Chng
Biogenesis of the outer membrane (OM) in Gram-negative bacteria, which is essential for viability, requires the coordinated transport and assembly of proteins and lipids, including lipopolysaccharides (LPS) and phospholipids (PLs), into the membrane. While pathways for LPS and OM protein assembly are well-studied, how PLs are transported to and from the OM is not clear. Mechanisms that ensure OM stability and homeostasis are also unknown. The trans-envelope Tol-Pal complex, whose physiological role has remained elusive, is important for OM stability...
August 16, 2017: Molecular Microbiology
Christine Diethmaier, Ravi Chawla, Alexandra Canzoneri, Daniel B Kearns, Pushkar P Lele, David Dubnau
Bacillus subtilis flagella are not only required for locomotion but also act as sensors that monitor environmental changes. Although how the signal transmission takes place is poorly understood, it has been shown that flagella play an important role in surface sensing by transmitting a mechanical signal to control the DegS-DegU two-component system. Here we report a role for flagella in the regulation of the K-state, which enables transformability and antibiotic tolerance (persistence). Mutations impairing flagellar synthesis are inferred to increase DegU-P, which inhibits the expression of ComK, the master regulator for the K-state, and reduces transformability...
August 11, 2017: Molecular Microbiology
Xin Zheng, Na Feng, Defeng Li, Xiuzhu Dong, Jie Li
RNase J, a prokaryotic 5'-3' exo/endoribonuclease, contributes to mRNA decay, rRNA maturation, and post-transcriptional regulation. Yet the processive-exoribonucleolysis mechanism remains obscure. Here, we solved the first RNA-free and RNA-bound structures of an archaeal RNase J, and through intensive biochemical studies provided detailed mechanistic insights into the catalysis and processivity. Distinct dimerization/tetramerization patterns were observed for archaeal and bacterial RNase Js, and unique archaeal Loops I and II were found involved in RNA interaction...
August 10, 2017: Molecular Microbiology
Bastian Groitl, Jan-Ulrik Dahl, Jeremy W Schroeder, Ursula Jakob
The most abundant oxidants controlling bacterial colonization on mucosal barrier epithelia are hypochlorous acid (HOCl), hypobromous acid (HOBr), and hypothiocyanous acid (HOSCN). All three oxidants are highly antimicrobial but little is known about their relative efficacies, their respective cellular targets, or what specific responses they elicit in bacteria. To address these important questions, we directly tested the individual oxidants on the virulent Pseudomonas aeruginosa strain PA14. We discovered that HOCl and HOBr work almost interchangeably, impacting non-growing bacterial cultures more significantly than actively growing bacteria, and eliciting similar stress responses, including the heat shock response...
August 10, 2017: Molecular Microbiology
Ciro D Cordeiro, Adolfo Saiardi, Roberto Docampo
Inositol pyrophosphates are novel signaling molecules possessing high-energy pyrophosphate bonds and involved in a number of biological functions. Here, we report the correct identification and characterization of the kinases involved in the inositol pyrophosphate biosynthetic pathway in Trypanosoma brucei: inositol polyphosphate multikinase (TbIPMK), inositol pentakisphosphate 2-kinase (TbIP5K) and inositol hexakisphosphate kinase (TbIP6K). TbIP5K and TbIP6K were not identifiable by sequence alone and their activities were validated by enzymatic assays with the recombinant proteins or by their complementation of yeast mutants...
August 9, 2017: Molecular Microbiology
Emma Louise Meredith, Ambika Kumar, Aya Konno, Joanna Szular, Sam Alsford, Karin Seifert, David Horn, Shane R Wilkinson
Quinone-based compounds have been exploited to treat infectious diseases and cancer, with such chemicals often functioning as inhibitors of key metabolic pathways or as prodrugs. Here, we screened an aziridinyl-1,4-benzoquinone (ABQ) library against the causative agents of trypanosomiasis, and cutaneous leishmaniasis, identifying several potent structures that exhibited EC50 values of <100 nM. However, these compounds also displayed significant toxicity towards mammalian cells indicating that they are not suitable therapies for systemic infections...
August 9, 2017: Molecular Microbiology
Jad Sassine, Meizhu Xu, Karzan R Sidiq, Robyn Emmins, Jeff Errington, Richard A Daniel
Bacterial cell division involves the dynamic assembly of a diverse set of proteins that coordinate the invagination of the cell membrane and synthesis of cell wall material to create the new cell poles of the separated daughter cells. Pencillin-binding protein PBP 2B is a key cell division protein in Bacillus subtilis proposed to have a specific catalytic role in septal wall synthesis. Unexpectedly, we find that a catalytically inactive mutant of PBP 2B supports cell division, but in this background the normally dispensable PBP 3 becomes essential...
August 9, 2017: Molecular Microbiology
Katherine R Hummels, Anne Witzky, Andrei Rajkovic, Rodney Tollerson, Lisa A Jones, Michael Ibba, Daniel B Kearns
Translation elongation factor P (EF-P) in Bacillus subtilis is required for a form of surface migration called swarming motility. Furthermore, B. subtilis EF-P is post-translationally modified with a 5-aminopentanol group but the pathway necessary for the synthesis and ligation of the modification is unknown. Here we determine that the protein YmfI catalyzes the reduction of EF-P-5 aminopentanone to EF-P-5 aminopentanol. In the absence of YmfI, accumulation of 5-aminopentanonylated EF-P is inhibitory to swarming motility...
August 8, 2017: Molecular Microbiology
Elyse E Muñoz, Kevin J Hart, Michael P Walker, Mark F Kennedy, Mackenzie M Shipley, Scott E Lindner
Transmission of the malaria parasite occurs in an unpredictable moment, when a mosquito takes a blood meal. Plasmodium has therefore evolved strategies to prepare for transmission, including translationally repressing and protecting mRNAs needed to establish the infection. However, mechanisms underlying these critical controls are not well understood, including whether Plasmodium changes its translationally repressive complexes and mRNA targets in different stages. Efforts to understand this have been stymied by severe technical limitations due to substantial mosquito contamination of samples...
August 8, 2017: Molecular Microbiology
Ying Zhang, Xiaolei Gao, Raphael Manck, Marjorie Schmid, Aysha H Osmani, Stephen A Osmani, Norio Takeshita, Reinhard Fischer
Microtubule-organizing centers (MTOCs) are large, multi-subunit protein complexes. Schizosaccharomyces pombe harbors MTOCs at spindle pole bodies, transient MTOCs in the division plane (eMTOCs) and nuclear-envelope associated MTOCs in interphase cells (iMTOCs). In the filamentous fungus Aspergillus nidulans SPBs and septum-associated MTOCs were described. Although comparable to S. pombe eMTOCs, A. nidulans sMTOCS are permanent septum-associated structures. The composition of sMTOCs is poorly understood and how they are targeted to septa was unknown...
August 8, 2017: Molecular Microbiology
Maike Bartholomae, Andrius Buivydas, Jakob H Viel, Manuel Montalban-Lopez, Oscar P Kuipers
Posttranslationally modified peptides commonly display antimicrobial activity, but can also aid the development of bacterial colonies, giving a competitive advantage in the ecological niche. The production of posttranslationally modified peptides by bacteria is a complex and energetically costly process that is strictly orchestrated in the cell. The onset of peptide production is linked to the different enzymes that take part during maturation, the transporters and the immunity determinants (if required). Thus, the population can make optimal use of available resources and obtain the benefits of production at an advantageous moment during growth, avoiding toxicity to itself...
August 8, 2017: Molecular Microbiology
Ye Won Cho, Alfonso Gonzales, Thomas V Harwood, Jessica Huynh, Yeji Hwang, Jun Sang Park, Anthony Q Trieu, Parth Italia, Vivek K Pallipuram, Douglas D Risser
Many cyanobacteria exhibit surface motility powered by type 4 pili (T4P). In the model filamentous cyanobacterium Nostoc punctiforme, the T4P systems are arrayed in static, bipolar rings in each cell. The chemotaxis-like Hmp system is essential for motility and the coordinated polar accumulation of PilA on cells in motile filaments, while the Ptx system controls positive phototaxis. Using transposon mutagenesis, a gene, designated hmpF, was identified as involved in motility. Synteny among filamentous cyanobacteria and the similar expression patterns for hmpF and hmpD imply that HmpF is part of the Hmp system...
August 5, 2017: Molecular Microbiology
Cecilia B Di Capua, Mariana Doprado, Juan Manuel Belardinelli, Héctor R Morbidoni
The synthesis of unsaturated fatty acids in Mycobacterium smegmatis is poorly characterized. Bioinformatic analysis revealed four putative fatty acid desaturases in its genome, one of which, MSMEG_1886, is highly homologous to desA3, the only palmitoyl/stearoyl desaturase present in the Mycobacterium tuberculosis genome. A MSMEG_1886 deletion mutant was partially auxotrophic for oleic acid and viable at 37°C and 25°C, although with a long lag phase in liquid medium. Fatty acid analysis suggested that MSMEG_1886 is a palmitoyl/stearoyl desaturase, as the synthesis of palmitoleic acid was abrogated, while oleic acid contents dropped by half in the mutant...
August 1, 2017: Molecular Microbiology
Morgan E Milton, C Leigh Allen, Erik A Feldmann, Benjamin G Bobay, David K Jung, Matthew D Stephens, Roberta J Melander, Kelly E Theisen, Daina Zeng, Richele J Thompson, Christian Melander, John Cavanagh
With antibiotic resistance increasing at alarming rates, targets for new antimicrobial therapies must be identified. A particularly promising target is the bacterial two-component system. Two-component systems allow bacteria to detect, evaluate and protect themselves against changes in the environment, such as exposure to antibiotics, and also to trigger production of virulence factors. Drugs that target the response regulator portion of two-component systems represent a potent new approach so far unexploited...
July 29, 2017: Molecular Microbiology
Virginia Basso, Sadri Znaidi, Valentine Lagage, Vitor Cabral, Franziska Schoenherr, Salomé LeibundGut-Landmann, Christophe d'Enfert, Sophie Bachellier-Bassi
Skn7 is a conserved fungal heat shock factor-type transcriptional regulator. It participates in maintaining cell wall integrity and regulates the osmotic/oxidative stress response (OSR) in S. cerevisiae, where it is part of a two-component signal transduction system. Here, we comprehensively address the function of Skn7 in the human fungal pathogen Candida albicans. We provide evidence reinforcing functional divergence, with loss of the cell wall/osmotic stress-protective roles and acquisition of the ability to regulate morphogenesis on solid medium...
July 28, 2017: Molecular Microbiology
Tracy A Washington, Janet L Smith, Alan D Grossman
DnaA is the widely conserved bacterial AAA+ ATPase that functions as both the replication initiator and a transcription factor. In many organisms, DnaA controls expression of its own gene and likely several others during growth and in response to replication stress. To evaluate the effects of DnaA on gene expression, separate from its role in replication initiation, we analyzed changes in mRNA levels in Bacillus subtilis cells with and without dnaA, using engineered strains in which dnaA is not essential. We found that dnaA was required for many of the changes in gene expression in response to replication stress...
July 27, 2017: Molecular Microbiology
Rebecca S Bamert, Karl Lundquist, Hyea Hwang, Chaille T Webb, Takoya Shiota, Christopher J Stubenrauch, Mathew J Belousoff, Robert J A Goode, Ralf B Schittenhelm, Richard Zimmerman, Martin Jung, James C Gumbart, Trevor Lithgow
The assembly of proteins into bacterial outer membranes is a key cellular process that we are only beginning to understand, mediated by the β-barrel assembly machinery (BAM). Two crucial elements of that machinery are the core BAM complex and the translocation and assembly module (TAM), with each containing a member of the Omp85 superfamily of proteins: BamA in the BAM complex, TamA in the TAM. Here, we used the substrate protein FimD as a model to assess the selectivity of substrate interactions for the TAM relative to those of the BAM complex...
July 27, 2017: Molecular Microbiology
Masafumi Kameya, Haruna Kanbe, Yasuo Igarashi, Hiroyuki Arai, Masaharu Ishii
Dissimilatory nitrate reductase (NAR) and assimilatory nitrate reductase (NAS) serve as key enzymes for nitrogen catabolism and anabolism in many organisms. We purified NAR and NAS from H. thermophilus, a hydrogen-oxidizing chemolithoautotroph belonging to the phylogenetically deepest branch in the Bacteria domain. Physiological contribution of these enzymes to nitrate respiration and assimilation was clarified by transcriptomic analysis and gene disruption experiments. These enzymes showed several features unreported in bacteria, such as the periplasmic orientation of NAR anchored with a putative transmembrane subunit and the specific electron transfer from a [4Fe-4S]-type ferredoxin to NAS...
July 27, 2017: Molecular Microbiology
Romain Christiano, Nikolay G Kolev, Huafang Shi, Elisabetta Ullu, Tobias C Walther, Christian Tschudi
The infectious metacyclic forms of Trypanosoma brucei result from a complex development in the tsetse fly vector. When they infect mammals, they cause African sleeping sickness in humans. Due to scarcity of biological material and difficulties of the tsetse fly as an experimental system, very limited information is available concerning the gene expression profile of metacyclic forms. We used an in vitro system based on expressing the RNA binding protein 6 to obtain infectious metacyclics and determined their protein and mRNA repertoires by mass-spectrometry (MS) based proteomics and mRNA sequencing (RNA-Seq) in comparison to non-infectious procyclic trypanosomes...
July 25, 2017: Molecular Microbiology
Markus Mörk-Mörkenstein, Ralf Heermann, Yvonne Göpel, Kirsten Jung, Boris Görke
The two-component system KdpD/KdpE governs K(+) homeostasis by controlling synthesis of the high affinity K(+) transporter KdpFABC. When sensing low environmental K(+) concentrations, the dimeric kinase KdpD autophosphorylates in trans and transfers the phosphoryl-group to the response regulator KdpE, which subsequently activates kdpFABC transcription. In Escherichia coli, KdpD can also be activated by interaction with the non-phosphorylated form of the accessory protein PtsN. PtsN stimulates KdpD kinase activity thereby increasing phospho-KdpE levels...
July 17, 2017: Molecular Microbiology
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