Characterization of the self-targeting Type IV CRISPR interference system in Pseudomonas oleovorans.

Bacterial Type IV CRISPR-Cas systems are thought to rely on multi-subunit ribonucleoprotein complexes to interfere with mobile genetic elements, but the substrate requirements and potential DNA nuclease activities for many systems within this type are uncharacterized. Here we show that the native Pseudomonas oleovorans Type IV-A CRISPR-Cas system targets DNA in a PAM-dependent manner and elicits interference without showing DNA nuclease activity. We found that the first crRNA of P.

Insights Into the Helical Shape Complex of .

One important factor that promotes the colonization of the upper digestive system of the human pathogen is its helical cell shape. The bacteria cell shape is predominantly defined by its peptidoglycan cell wall. In rod-shaped species, PG synthesis is mediated by two dynamic molecular machines that facilitate growth along the perpendicular axis and the septum, called the elongasome and the divisome, respectively.

Intersubunit Coupling Enables Fast CO-Fixation by Reductive Carboxylases.

Enoyl-CoA carboxylases/reductases (ECRs) are some of the most efficient CO-fixing enzymes described to date. However, the molecular mechanisms underlying the extraordinary catalytic activity of ECRs on the level of the protein assembly remain elusive. Here we used a combination of ambient-temperature X-ray free electron laser (XFEL) and cryogenic synchrotron experiments to study the structural organization of the ECR from .

Spatiotemporal Coupling of DNA Supercoiling and Genomic Sequence Organization-A Timing Chain for the Bacterial Growth Cycle?

In this article we describe the bacterial growth cycle as a closed, self-reproducing, or autopoietic circuit, reestablishing the physiological state of stationary cells initially inoculated in the growth medium. In batch culture, this process of self-reproduction is associated with the gradual decline in available metabolic energy and corresponding change in the physiological state of the population as a function of "travelled distance" along the autopoietic path. We argue that this directional alteration of cell physiology is both reflected in and supported by sequential gene expression along the chromosomal OriC-Ter axis.

Hippocampal Cytokine Release in Experimental Epileptogenesis-A Longitudinal In Vivo Microdialysis Study.

Background: Inflammation, particularly cytokine release, contributes to epileptogenesis by influencing the cerebral tissue remodeling and neuronal excitability that occurs after a precipitating epileptogenic insult. While several cytokines have been explored in this process, release kinetics are less well investigated. Determining the time course of cytokine release in the epileptogenic zone is necessary for precisely timed preventive or therapeutic anti-inflammatory interventions.

Rocaglates as Antivirals: Comparing the Effects on Viral Resistance, Anti-Coronaviral Activity, RNA-Clamping on eIF4A and Immune Cell Toxicity.

Rocaglates are potent broad-spectrum antiviral compounds with a promising safety profile. They inhibit viral protein synthesis for different RNA viruses by clamping the 5'-UTRs of mRNAs onto the surface of the RNA helicase eIF4A. Apart from the natural rocaglate silvestrol, synthetic rocaglates like zotatifin or CR-1-31-B have been developed.

Discovery of a New Chemoeffector for Chemoreceptor Tsr and Identification of a Molecular Mechanism of Repellent Sensing.

Motile bacteria use chemotaxis to search for nutrients and escape from harmful chemicals. While the sensing mechanisms for chemical attractants are well established, the molecular details of chemorepellent detection are poorly understood. Here, by using combined computational and experimental approaches to screen potential chemoeffectors for the chemoreceptor Tsr, we identified a specific chemorepellent, 1-aminocyclohexanecarboxylic acid (ACHC).

In Vitro Activity of Amphotericin B in Combination with Colistin against Fungi Responsible for Invasive Infections.

The in vitro interaction of amphotericin B in combination with colistin was evaluated against a total of 86 strains comprising of 47 species (10 , 15 , five , three , five , four and five ), 29 species (five , 10 , four , five , and five ), and 10 species (seven , one and two ) strains. For the determination of the interaction, a microdilution checkerboard technique based on the European Committee on Antimicrobial Susceptibility Testing (EUCAST) reference method for antifungal susceptibility testing was used. Results of the checkerboard technique were evaluated by the fractional inhibitory concentration index (FICI) based on the Loewe additivity model for all isolates.

An overview of bioinformatics, genomics, and transcriptomics resources for bryophytes.

Bryophytes are useful models for the study of plant evolution, development, plant-fungal symbiosis, stress responses, and gametogenesis. Additionally, their dominant haploid gametophytic phase makes them great models for functional genomics research, allowing straightforward genome editing and gene knockout via CRISPR or homologous recombination. Until 2016, however, the only bryophyte genome sequence published was that of Physcomitrium patens.

The quantitative basis for the redistribution of immobile bacterial lipoproteins to division septa.

The spatial localisation of proteins is critical for most cellular function. In bacteria, this is typically achieved through capture by established landmark proteins. However, this requires that the protein is diffusive on the appropriate timescale.

Optogenetics in bacteria - applications and opportunities.

Optogenetics holds the promise of controlling biological processes with superb temporal and spatial resolution at minimal perturbation. Although many of the light-reactive proteins used in optogenetic systems are derived from prokaryotes, applications were largely limited to eukaryotes for a long time. In recent years, however, an increasing number of microbiologists use optogenetics as a powerful new tool to study and control key aspects of bacterial biology in a fast and often reversible manner.

Bacterial Small Membrane Proteins: the Swiss Army Knife of Regulators at the Lipid Bilayer.

Small membrane proteins represent a subset of recently discovered small proteins (≤100 amino acids), which are a ubiquitous class of emerging regulators underlying bacterial adaptation to environmental stressors. Until relatively recently, small open reading frames encoding these proteins were not designated genes in genome annotations. Therefore, our understanding of small protein biology was primarily limited to a few candidates associated with previously characterized larger partner proteins.

Transcription-dependent confined diffusion of enzymes within subcellular spaces of the bacterial cytoplasm.

Background: Knowledge on the localization and mobility of enzymes inside bacterial cells is scarce, but important for understanding spatial regulation of metabolism. The four central enzymes (Rib enzymes) of the riboflavin (RF) biosynthesis pathway in the Gram positive model bacterium Bacillus subtilis have been studied extensively in vitro, especially the heavy RF synthase, a large protein complex with a capsid structure formed by RibH and an encapsulated RibE homotrimer, which mediates substrate-channeling. However, little is known about the behavior and mobility of these enzymes in vivo.

Actin-generated force applied during endocytosis measured by Sla2-based FRET tension sensors.

Mechanical forces are integral to many cellular processes, including clathrin-mediated endocytosis, a principal membrane trafficking route into the cell. During endocytosis, forces provided by endocytic proteins and the polymerizing actin cytoskeleton reshape the plasma membrane into a vesicle. Assessing force requirements of endocytic membrane remodeling is essential for understanding endocytosis.

Change in Cofactor Specificity of Oxidoreductases by Adaptive Evolution of an Escherichia coli NADPH-Auxotrophic Strain.

The nicotinamide cofactor specificity of enzymes plays a key role in regulating metabolic processes and attaining cellular homeostasis. Multiple studies have used enzyme engineering tools or a directed evolution approach to switch the cofactor preference of specific oxidoreductases. However, whole-cell adaptation toward the emergence of novel cofactor regeneration routes has not been previously explored.

Functional compartmentalization and metabolic separation in a prokaryotic cell.

The prokaryotic cell is traditionally seen as a "bag of enzymes," yet its organization is much more complex than in this simplified view. By now, various microcompartments encapsulating metabolic enzymes or pathways are known for These microcompartments are usually small, encapsulating and concentrating only a few enzymes, thus protecting the cell from toxic intermediates or preventing unwanted side reactions. The hyperthermophilic, strictly anaerobic Crenarchaeon is an extraordinary organism possessing two membranes, an inner and an energized outer membrane.

Asymmetric mating behavior of isogamous budding yeast.

Anisogamy, the size difference between small male and large female gametes, is known to enable selection for sexual dimorphism and behavioral differences between sexes. Nevertheless, even isogamous species exhibit molecular asymmetries between mating types, which are known to ensure their self-incompatibility. Here, we show that different properties of the pheromones secreted by the a and α mating types of budding yeast lead to asymmetry in their behavioral responses during mating in mixed haploid populations, which resemble behavioral asymmetries between gametes in anisogamous organisms.

Production and Characterization of Motile and Chemotactic Bacterial Minicells.

Minicells are nanosized membrane vesicles produced by bacteria. Minicells are chromosome-free but contain cellular biosynthetic and metabolic machinery, and they are robust due to the protection provided by the bacterial cell envelope, which makes them potentially highly attractive in biomedical applications. However, the applicability of minicells and other nanoparticle-based delivery systems is limited by their inefficient accumulation at the target.

Benzylmalonyl-CoA dehydrogenase, an enzyme involved in bacterial auxin degradation.

A novel acyl-CoA dehydrogenase involved in degradation of the auxin indoleacetate by Aromatoleum aromaticum was identified as a decarboxylating benzylmalonyl-CoA dehydrogenase (IaaF). It is encoded within the iaa operon coding for enzymes of indoleacetate catabolism. Using enzymatically produced benzylmalonyl-CoA, the reaction was characterized as simultaneous oxidation and decarboxylation of benzylmalonyl-CoA to cinnamoyl-CoA and CO.

Coregulation of gene expression by White collar 1 and phytochrome in Ustilago maydis.

Ustilago maydis encodes ten predicted light-sensing proteins. The biological functions of only a few of them are elucidated. Among the characterized ones are two DNA-photolyases and two rhodopsins that act as DNA-repair enzymes or green light-driven proton pumps, respectively.

Structure of the endocytic adaptor complex reveals the basis for efficient membrane anchoring during clathrin-mediated endocytosis.

During clathrin-mediated endocytosis, a complex and dynamic network of protein-membrane interactions cooperate to achieve membrane invagination. Throughout this process in yeast, endocytic coat adaptors, Sla2 and Ent1, must remain attached to the plasma membrane to transmit force from the actin cytoskeleton required for successful membrane invagination. Here, we present a cryo-EM structure of a 16-mer complex of the ANTH and ENTH membrane-binding domains from Sla2 and Ent1 bound to PIP that constitutes the anchor to the plasma membrane.

A synthetic C4 shuttle via the β-hydroxyaspartate cycle in C3 plants.

Plants depend on the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) for CO fixation. However, especially in C3 plants, photosynthetic yield is reduced by formation of 2-phosphoglycolate, a toxic oxygenation product of Rubisco, which needs to be recycled in a high-flux-demanding metabolic process called photorespiration. Canonical photorespiration dissipates energy and causes carbon and nitrogen losses.

Dynamics of Bacterial Signal Recognition Particle at a Single Molecule Level.

We have studied the localization and dynamics of bacterial Ffh, part of the SRP complex, its receptor FtsY, and of ribosomes in the Gamma-proteobacterium Using structured illumination microscopy, we show that ribosomes show a pronounced accumulation at the cell poles, whereas SRP and FtsY are distributed at distinct sites along the cell membrane, but they are not accumulated at the poles. Single molecule dynamics can be explained by assuming that all three proteins/complexes move as three distinguishable mobility fractions: a low mobility/static fraction may be engaged in translation, medium-fast diffusing fractions may be transition states, and high mobility populations likely represent freely diffusing molecules/complexes. Diffusion constants suggest that SRP and FtsY move together with slow-mobile ribosomes.