Microtubules provide force to promote membrane uncoating in vacuolar escape for a cyto-invasive bacterial pathogen.

Intracellular bacterial pathogens gain entry to mammalian cells inside a vacuole derived from the host membrane. Some of them escape the bacteria-containing vacuole (BCV) and colonize the cytosol. Bacteria replicating within BCVs coopt the microtubule network to position it within infected cells, whereas the role of microtubules for cyto-invasive pathogens remains obscure.

RACK1 promotes actin-mediated invasion, motility, and cell-to-cell spreading.

is an intracellular bacterium that hijacks the host actin cytoskeleton to invade and disseminate within the colonic epithelium. 's virulence factors induce actin polymerization, leading to bacterial uptake, actin tail formation, actin-mediated motility, and cell-to-cell spreading. Many host factors involved in the -prompted actin rearrangements remain elusive.

Expansion-assisted selective plane illumination microscopy for nanoscale imaging of centimeter-scale tissues.

Recent advances in tissue processing, labeling, and fluorescence microscopy are providing unprecedented views of the structure of cells and tissues at sub-diffraction resolutions and near single molecule sensitivity, driving discoveries in diverse fields of biology, including neuroscience. Biological tissue is organized over scales of nanometers to centimeters. Harnessing molecular imaging across intact, three-dimensional samples on this scale requires new types of microscopes with larger fields of view and working distance, as well as higher throughput.

Comparative study of GBP recruitment on two cytosol-dwelling pathogens, Francisella novicida and Shigella flexneri highlights differences in GBP repertoire and in GBP1 motif requirements.

Guanylate-Binding Proteins are interferon-inducible GTPases that play a key role in cell autonomous responses against intracellular pathogens. Despite sharing high sequence similarity, subtle differences among GBPs translate into functional divergences that are still largely not understood. A key GBP feature is the formation of supramolecular GBP complexes on the bacterial surface.

Fluorescence Lifetime Measurement of Prefibrillar Sickle Hemoglobin Oligomers as a Platform for Drug Discovery in Sickle Cell Disease.

The molecular origin of sickle cell disease (SCD) has been known since 1949, but treatments remain limited. We present the first high-throughput screening (HTS) platform for discovering small molecules that directly inhibit sickle hemoglobin (HbS) oligomerization and improve blood flow, potentially overcoming a long-standing bottleneck in SCD drug discovery. We show that at concentrations far below the threshold for nucleation and rapid polymerization, deoxygenated HbS forms small assemblies of multiple αβ tetramers.

Lambda Red Recombineering in Shigella flexneri.

Shigellosis remains a major cause of severe diarrheal disease and death throughout the world. Vaccine development against shigellosis has been hampered by an incomplete understanding of the molecular mechanisms by which Shigella spp. causes disease and difficulties in manipulating Shigella spp.

The Functional Differences between the GroEL Chaperonin of and the HtpB Chaperonin of Can Be Mapped to Specific Amino Acid Residues.

Group I chaperonins are a highly conserved family of essential proteins that self-assemble into molecular nanoboxes that mediate the folding of cytoplasmic proteins in bacteria and organelles. GroEL, the chaperonin of , is the archetype of the family. Protein folding-independent functions have been described for numerous chaperonins, including HtpB, the chaperonin of the bacterial pathogen .

TORC1 signaling modulates Cdk8-dependent GAL gene expression in Saccharomyces cerevisiae.

Cdk8 of the RNA polymerase II mediator kinase complex regulates gene expression by phosphorylating sequence-specific transcription factors. This function is conserved amongst eukaryotes, but the signals and mechanisms regulating Cdk8 activity and phosphorylation of its substrates are unknown. Full induction of the GAL genes in yeast requires phosphorylation of the transcriptional activator Gal4 by Cdk8.

Unique factors to the implementation of a disaster preparedness plan at Georgetown Public Hospital in Guyana.

Disaster planning in developing countries is largely ineffective despite these countries being more likely to suffer from catastrophic events. This article aims to identify strengths, educational needs, practice improvement opportunities, and local factors that may contribute to the development/implementation of a disaster preparedness plan at Georgetown Public Hospital in Guyana.

FRET and optical trapping reveal mechanisms of actin activation of the power stroke and phosphate release in myosin V.

Myosins generate force and motion by precisely coordinating their mechanical and chemical cycles, but the nature and timing of this coordination remains controversial. We utilized a FRET approach to examine the kinetics of structural changes in the force-generating lever arm in myosin V. We directly compared the FRET results with single-molecule mechanical events examined by optical trapping.

Guanylate-binding proteins convert cytosolic bacteria into caspase-4 signaling platforms.

Bacterial lipopolysaccharide triggers human caspase-4 (murine caspase-11) to cleave gasdermin-D and induce pyroptotic cell death. How lipopolysaccharide sequestered in the membranes of cytosol-invading bacteria activates caspases remains unknown. Here we show that in interferon-γ-stimulated cells guanylate-binding proteins (GBPs) assemble on the surface of Gram-negative bacteria into polyvalent signaling platforms required for activation of caspase-4.

Disruption of the extracellular polymeric network of Pseudomonas aeruginosa biofilms by alginate lyase enhances pathogen eradication by antibiotics.

Background: Pseudomonas aeruginosa forms antibiotic-resistant biofilms that are responsible for the treatment failure or relapses of the bacterial infections in the lungs of patients with cystic fibrosis (CF). The alginate lyases that target extracellular polysaccharide alginate of P. aeruginosa biofilms are promising therapeutic candidates for treatment of P.

Recalls of Foods due to Microbial Contamination Classified by the Canadian Food Inspection Agency, 2000 to 2017.

Recall of microbial-contaminated food products is an important intervention in preventing the transmission of foodborne illness. Here, we summarize the number and nature of foods recalled as a result of microbial contamination, classified by the Canadian Food Inspection Agency, for the period 1 January 2000 through 31 December 2017. A total of 10,432 food products were recalled from 2,094 recall events in Canada because of microbial contamination during this period.

promotes major alteration of gut epithelial physiology and tissue invasion by shutting off host intracellular transport.

Intracellular trafficking pathways in eukaryotic cells are essential to maintain organelle identity and structure, and to regulate cell communication with its environment. invades and subverts the human colonic epithelium by the injection of virulence factors through a type 3 secretion system (T3SS). In this work, we report the multiple effects of two effectors, IpaJ and VirA, which target small GTPases of the Arf and Rab families, consequently inhibiting several intracellular trafficking pathways.

Marine Bacteria, A Source for Alginolytic Enzyme to Disrupt Biofilms.

biofilms are typically associated with the chronic lung infection of cystic fibrosis (CF) patients and represent a major challenge for treatment. This opportunistic bacterial pathogen secretes alginate, a polysaccharide that is one of the main components of its biofilm. Targeting this major biofilm component has emerged as a tempting therapeutic strategy for tackling biofilm-associated bacterial infections.

Converter domain mutations in myosin alter structural kinetics and motor function.

Myosins are molecular motors that use a conserved ATPase cycle to generate force. We investigated two mutations in the converter domain of myosin V (R712G and F750L) to examine how altering specific structural transitions in the motor ATPase cycle can impair myosin mechanochemistry. The corresponding mutations in the human β-cardiac myosin gene are associated with hypertrophic and dilated cardiomyopathy, respectively.

Mavacamten stabilizes an autoinhibited state of two-headed cardiac myosin.

We used transient biochemical and structural kinetics to elucidate the molecular mechanism of mavacamten, an allosteric cardiac myosin inhibitor and a prospective treatment for hypertrophic cardiomyopathy. We find that mavacamten stabilizes an autoinhibited state of two-headed cardiac myosin not found in the single-headed S1 myosin motor fragment. We determined this by measuring cardiac myosin actin-activated and actin-independent ATPase and single-ATP turnover kinetics.

Complement C3 Drives Autophagy-Dependent Restriction of Cyto-invasive Bacteria.

In physiological settings, the complement protein C3 is deposited on all bacteria, including invasive pathogens. However, because experimental host-bacteria systems typically use decomplemented serum to avoid the lytic action of complement, the impact of C3 coating on epithelial cell responses to invasive bacteria remains unexplored. Here, we demonstrate that following invasion, intracellular C3-positive Listeria monocytogenes is targeted by autophagy through a direct C3/ATG16L1 interaction, resulting in autophagy-dependent bacterial growth restriction.

GBPs Inhibit Motility of Shigella flexneri but Are Targeted for Degradation by the Bacterial Ubiquitin Ligase IpaH9.8.

Interferon exposure boosts cell-autonomous immunity for more efficient pathogen control. But how interferon-enhanced immunity protects the cytosol against bacteria and how professionally cytosol-dwelling bacteria avoid clearance are insufficiently understood. Here we demonstrate that the interferon-induced GTPase family of guanylate-binding proteins (GBPs) coats Shigella flexneri in a hierarchical manner reliant on GBP1.

A Cardiomyopathy Mutation in the Myosin Essential Light Chain Alters Actomyosin Structure.

We have used site-directed time-resolved fluorescence resonance energy transfer to determine the effect of a pathological mutation in the human ventricular essential light chain (hVELC) of myosin, on the structural dynamics of the actin-myosin complex. The hVELC modulates the function of actomyosin, through the interaction of its N-terminal extension with actin and its C-terminal lobe with the myosin heavy chain. Several mutations in hVELC are associated with hypertrophic cardiomyopathy (HCM).