Functionality of Two Origins of Replication in Strains With a Single Chromosome.

Chromosomal inheritance in bacteria usually entails bidirectional replication of a single chromosome from a single origin into two copies and subsequent partitioning of one copy each into daughter cells upon cell division. However, the human pathogen and other harbor two chromosomes, a large Chr1 and a small Chr2. Chr1 and Chr2 have different origins, an type origin and a P1 plasmid-type origin, respectively, driving the replication of respective chromosomes.

Single molecule tracking reveals spatio-temporal dynamics of bacterial DNA repair centres.

Single-particle (molecule) tracking (SPT/SMT) is a powerful method to study dynamic processes in living bacterial cells at high spatial and temporal resolution. We have performed single-molecule imaging of early DNA double-strand break (DSB) repair events during homologous recombination in the model bacterium Bacillus subtilis. Our findings reveal that DNA repair centres arise at all sites on the chromosome and that RecN, RecO and RecJ perform fast, enzyme-like functions during detection and procession of DNA double strand breaks, respectively.

Type IV CRISPR RNA processing and effector complex formation in Aromatoleum aromaticum.

Type IV CRISPR-Cas modules belong to class 1 prokaryotic adaptive immune systems, which are defined by the presence of multisubunit effector complexes. They usually lack the known Cas proteins involved in adaptation and target cleavage, and their function has not been experimentally addressed. To investigate RNA and protein components of this CRISPR-Cas type, we located a complete type IV cas gene locus and an adjacent CRISPR array on a megaplasmid of Aromatoleum aromaticum EbN1, which contains an additional type I-C system on its chromosome.

The multicatalytic compartment of propionyl-CoA synthase sequesters a toxic metabolite.

Cells must cope with toxic or reactive intermediates formed during metabolism. One coping strategy is to sequester reactions that produce such intermediates within specialized compartments or tunnels connecting different active sites. Here, we show that propionyl-CoA synthase (PCS), an ∼ 400-kDa homodimer, three-domain fusion protein and the key enzyme of the 3-hydroxypropionate bi-cycle for CO fixation, sequesters its reactive intermediate acrylyl-CoA.

Structural basis for (p)ppGpp-mediated inhibition of the GTPase RbgA.

Efficient adaptation to environmental changes is pivotal for all bacterial cells. Almost all bacterial species depend on the conserved stringent response system to prompt timely transcriptional and metabolic responses according to stress conditions and nutrient depletion. The stringent response relies on the stress-dependent synthesis of the second messenger nucleotides and alarmones (p)ppGpp, which pleiotropically target and reprogram processes that consume cellular resources, such as ribosome biogenesis.

SMTracker: a tool for quantitative analysis, exploration and visualization of single-molecule tracking data reveals highly dynamic binding of B. subtilis global repressor AbrB throughout the genome.

Single-particle (molecule) tracking (SPT/SMT) is a powerful method to study dynamic processes in living cells at high spatial and temporal resolution. Even though SMT is becoming a widely used method in bacterial cell biology, there is no program employing different analytical tools for the quantitative evaluation of tracking data. We developed SMTracker, a MATLAB-based graphical user interface (GUI) for automatically quantifying, visualizing and managing SMT data via five interactive panels, allowing the user to interactively explore tracking data from several conditions, movies and cells on a track-by-track basis.

A System for Gene Expression Noise Control in Yeast.

Gene expression noise arises from stochastic variation in the synthesis and degradation of mRNA and protein molecules and creates differences in protein numbers across populations of genetically identical cells. Such variability can lead to imprecision and reduced performance of both native and synthetic networks. In principle, gene expression noise can be controlled through the rates of transcription, translation and degradation, such that different combinations of those rates lead to the same protein concentrations but at different noise levels.

The germinal centre kinase Don3 is crucial for unconventional secretion of chitinase Cts1 in Ustilago maydis.

Unconventional secretion has emerged as an increasingly important cellular process in eukaryotic cells. The underlying translocation mechanisms are diverse and often little understood. We study unconventional secretion of chitinase Cts1 in the corn smut fungus Ustilago maydis.

'Negative' and 'positive catalysis': complementary principles that shape the catalytic landscape of enzymes.

Our understanding of enzyme catalysis is dominated by transition state theory. According to this concept, an enzymatic reaction is guided along a desired reaction coordinate through the stabilization of favorable transition state. But how much is the outcome of an enzyme reaction controlled by the destabilization of unwanted transition states? Here, we revive and critically review the hypothesis that the active site of enzymes also features elements of 'negative catalysis'.

Chromate Resistance Mechanisms in Leucobacter chromiiresistens.

Chromate is one of the major anthropogenic contaminants on Earth. is a highly chromate-resistant strain, tolerating chromate concentrations in LB medium of up to 400 mM. In response to chromate stress, forms biofilms, which are held together via extracellular DNA.

statTarget: A streamlined tool for signal drift correction and interpretations of quantitative mass spectrometry-based omics data.

Large-scale quantitative mass spectrometry-based metabolomics and proteomics study requires the long-term analysis of multiple batches of biological samples, which often accompanied with significant signal drift and various inter- and intra-batch variations. The unwanted variations can lead to poor inter- and intra-day reproducibility, which is a hindrance to discover real significance. The use of quality control samples and data treatment strategies in the quality assurance procedure provides a mechanism to evaluate the quality and remove the analytical variance of the data.

Identification and Localization of Peroxisomal Biogenesis Proteins Indicates the Presence of Peroxisomes in the Cryptophyte Guillardia theta and Other "Chromalveolates".

Peroxisomes are single-membrane-bound organelles with a huge metabolic versatility, including the degradation of fatty acids (β-oxidation) and the detoxification of reactive oxygen species as most conserved functions. Although peroxisomes seem to be present in the majority of investigated eukaryotes, where they are responsible for many eclectic and important spatially separated metabolic reactions, knowledge about their existence in the plethora of protists (eukaryotic microorganisms) is scarce. Here, we investigated genomic data of organisms containing complex plastids with red algal ancestry (so-called "chromalveolates") for the presence of genes encoding peroxins-factors specific for the biogenesis, maintenance, and division of peroxisomes in eukaryotic cells.

Unique Archaeal Small RNAs.

Advances in genome-wide sequence technologies allow for detailed insights into the complexity of RNA landscapes of organisms from all three domains of life. Recent analyses of archaeal transcriptomes identified interaction and regulation networks of noncoding RNAs in this understudied domain. Here, we review current knowledge of small, noncoding RNAs with important functions for the archaeal lifestyle, which often requires adaptation to extreme environments.

Microdomain formation is a general property of bacterial membrane proteins and induces heterogeneity of diffusion patterns.

Background: Proteins within the cytoplasmic membrane display distinct localization patterns and arrangements. While multiple models exist describing the dynamics of membrane proteins, to date, there have been few systematic studies, particularly in bacteria, to evaluate how protein size, number of transmembrane domains, and temperature affect their diffusion, and if conserved localization patterns exist.

Results: We have used fluorescence microscopy, single-molecule tracking (SMT), and computer-aided visualization methods to obtain a better understanding of the three-dimensional organization of bacterial membrane proteins, using the model bacterium Bacillus subtilis.

PAM identification by CRISPR-Cas effector complexes: diversified mechanisms and structures.

Adaptive immunity of prokaryotes is mediated by CRISPR-Cas systems that employ a large variety of Cas protein effectors to identify and destroy foreign genetic material. The different targeting mechanisms of Cas proteins rely on the proper protection of the host genome sequence while allowing for efficient detection of target sequences, termed protospacers. A short DNA sequence, the protospacer-adjacent motif (PAM), is frequently used to mark proper target sites.

Modular Assembly of Synthetic Secondary Chromosomes.

The development of novel DNA assembly methods in recent years has paved the way for the construction of synthetic replicons to be used for basic research and biotechnological applications. Questions of how chromosomes need to be constructed to maintain the genetic information can now be answered by a learning-by-building approach. Here, we describe an efficient pipeline for the design and assembly of synthetic, secondary chromosomes in Escherichia coli based on the popular Modular Cloning system (MoClo).

Seven-transmembrane receptor protein RgsP and cell wall-binding protein RgsM promote unipolar growth in Rhizobiales.

Members of the Rhizobiales (class of α-proteobacteria) display zonal peptidoglycan cell wall growth at one cell pole, contrasting with the dispersed mode of cell wall growth along the sidewalls of many other rod-shaped bacteria. Here we show that the seven-transmembrane receptor (7TMR) protein RgsP (SMc00074), together with the putative membrane-anchored peptidoglycan metallopeptidase RgsM (SMc02432), have key roles in unipolar peptidoglycan formation during growth and at mid-cell during cell division in Sinorhizobium meliloti. RgsP is composed of a periplasmic globular 7TMR-DISMED2 domain, a membrane-spanning region, and cytoplasmic PAS, GGDEF and EAL domains.

Iron-Sulfur Cluster Biosynthesis in Algae with Complex Plastids.

Plastids surrounded by four membranes harbor a special compartment between the outer and inner plastid membrane pair, the so-called periplastidal compartment (PPC). This cellular structure is usually presumed to be the reduced cytoplasm of a eukaryotic phototrophic endosymbiont, which was integrated into a host cell and streamlined into a plastid with a complex membrane structure. Up to date, no mitochondrion or mitochondrion-related organelle has been identified in the PPC of any representative.

FliS/flagellin/FliW heterotrimer couples type III secretion and flagellin homeostasis.

Flagellin is amongst the most abundant proteins in flagellated bacterial species and constitutes the major building block of the flagellar filament. The proteins FliW and FliS serve in the post-transcriptional control of flagellin and guide the protein to the flagellar type III secretion system (fT3SS), respectively. Here, we present the high-resolution structure of FliS/flagellin heterodimer and show that FliS and FliW bind to opposing interfaces located at the N- and C-termini of flagellin.

Inverted signaling by bacterial chemotaxis receptors.

Microorganisms use transmembrane sensory receptors to perceive a wide range of environmental factors. It is unclear how rapidly the sensory properties of these receptors can be modified when microorganisms adapt to novel environments. Here, we demonstrate experimentally that the response of an Escherichia coli chemotaxis receptor to its chemical ligands can be easily inverted by mutations at several sites along receptor sequence.

Crude-MS Strategy for in-Depth Proteome Analysis of the Methane-Oxidizing Methylocystis sp. strain SC2.

Methylocystis sp. strain SC2 is a representative of the alphaproteobacterial methane oxidizers or type IIa methanotrophs. These microorganisms play a crucial role in methane cycling.

A gated relaxation oscillator mediated by FrzX controls morphogenetic movements in Myxococcus xanthus.

Dynamic control of cell polarity is of critical importance for many aspects of cellular development and motility. In Myxococcus xanthus, MglA, a G protein, and MglB, its cognate GTPase-activating protein, establish a polarity axis that defines the direction of movement of the cell and that can be rapidly inverted by the Frz chemosensory system. Although vital for collective cell behaviours, how Frz triggers this switch has remained unknown.

A novel complex neurological phenotype due to a homozygous mutation in FDX2.

Defects in iron-sulphur [Fe-S] cluster biogenesis are increasingly recognized as causing neurological disease. Mutations in a number of genes that encode proteins involved in mitochondrial [Fe-S] protein assembly lead to complex neurological phenotypes. One class of proteins essential in the early cluster assembly are ferredoxins.

ZomB is essential for flagellar motor reversals in Shewanella putrefaciens and Vibrio parahaemolyticus.

The ability of most bacterial flagellar motors to reverse the direction of rotation is crucial for efficient chemotaxis. In Escherichia coli, motor reversals are mediated by binding of phosphorylated chemotaxis protein CheY to components of the flagellar rotor, FliM and FliN, which induces a conformational switch of the flagellar C-ring. Here, we show that for Shewanella putrefaciens, Vibrio parahaemolyticus and likely a number of other species an additional transmembrane protein, ZomB, is critically required for motor reversals as mutants lacking ZomB exclusively exhibit straightforward swimming also upon full phosphorylation or overproduction of CheY.

Chemotaxis of Escherichia coli to major hormones and polyamines present in human gut.

The microorganisms in the gastrointestinal (GI) tract can influence the metabolism, immunity, and behavior of animal hosts. Increasing evidence suggests that communication between the host and the microbiome also occurs in the opposite direction, with hormones and other host-secreted compounds being sensed by microorganisms. Here, we addressed one key aspect of the host-microbe communication by studying chemotaxis of a model commensal bacterium, Escherichia coli, to several compounds present abundantly in the GI tract, namely catecholamines, thyroid hormones, and polyamines.