Malaria parasite egress at a glance

ABSTRACTAll intracellular pathogens must escape (egress) from the confines of their host cell to disseminate and proliferate. The malaria parasite only replicates in an intracellular vacuole or in a cyst, and must undergo egress at four distinct phases during its complex life cycle, each time disrupting, in a highly regulated manner, the membranes or cyst wall that entrap the parasites. This Cell Science at a Glance article and accompanying poster summarises our current knowledge of the morphological features of egress across the Plasmodium life cycle, the molecular mechanisms that govern the process, and how researchers are working to exploit this knowledge to develop much-needed new approaches to malaria control.

Inactivation of mediator complex protein 22 in podocytes results in intracellular vacuole formation, podocyte loss and premature death

Podocytes are critical for the maintenance of kidney ultrafiltration barrier and play a key role in the progression of glomerular diseases. Although mediator complex proteins have been shown to be important for many physiological and pathological processes, their role in kidney tissue has not been studied. In this study, we identified a mediator complex protein 22 (Med22) as a renal podocyte cell-enriched molecule. Podocyte-specific Med22 knockout mouse showed that Med22 was not needed for normal podocyte maturation. However, it was critical for the maintenance of podocyte health as the mice developed progressive glomerular disease and died due to renal failure. Detailed morphological analyses showed that Med22-deficiency in podocytes resulted in intracellular vacuole formation followed by podocyte loss. Moreover, Med22-deficiency in younger mice promoted the progression of glomerular disease, suggesting Med22-mediated processes may have a role in the development of glomerulopathies. This study shows for the first time that mediator complex has a critical role in kidney physiology.

Bacillus methylotrophicus has potential applications against Monilinia fructicola

Biocontrol is a cost-effective and environmentally friendly technique used in agricultural production. We isolated and screened a bacterial strain from the soils of a peach orchard with high yield. Using biochemical and physiological analysis as well as phylogenetic sequencing data, we identified a strain of Bacillus methylotrophicus, strain XJ-C. The results of our screening trials showed that XJ-C was able to suppress M. fructicola at an inhibition rate of 81.57%. Following the application of a 1×109 CFU/mL XJ-C strain suspension to the fruits, leaves, and shoots of peach trees infected with M. fructicola, the inhibition rate reached 64.31%, 97.34%, and 64.28%, respectively. Using OM and SEM, we observed that, under the inhibition of strain XJ-C, M. fructicola mycelium and spores were abnormally shaped. Under TEM, cell walls were transparent, organelles had disappeared, and the intracellular vacuole was deformed. Thus, XJ-C has the potential to be used in biocontrol.

The Cell Wall Lipid PDIM Contributes to Phagosomal Escape and Host Cell Exit of Mycobacterium tuberculosis

ABSTRACT The cell wall of Mycobacterium tuberculosis is composed of unique lipids that are important for pathogenesis. Indeed, the first-ever genetic screen in M. tuberculosis identified genes involved in the biosynthesis and transport of the cell wall lipid PDIM (phthiocerol dimycocerosates) as crucial for the survival of M. tuberculosis in mice. Here we show evidence for a novel molecular mechanism of the PDIM-mediated virulence in M. tuberculosis. We characterized the DNA interaction and the regulon of Rv3167c, a transcriptional repressor that is involved in virulence regulation of M. tuberculosis, and discovered that it controls the PDIM operon. A loss-of-function genetic approach showed that PDIM levels directly correlate with the capacity of M. tuberculosis to escape the phagosome and induce host cell necrosis and macroautophagy. In conclusion, our study attributes a novel role of the cell wall lipid PDIM in intracellular host cell modulation, which is important for host cell exit and dissemination of M. tuberculosis. IMPORTANCE Mycobacterium tuberculosis is a major human pathogen that has coevolved with its host for thousands of years. The complex and unique cell wall of M. tuberculosis contains the lipid PDIM (phthiocerol dimycocerosates), which is crucial for virulence of the bacterium, but its function is not well understood. Here we show that PDIM expression by M. tuberculosis is negatively regulated by a novel transcriptional repressor, Rv3167c. In addition, we discovered that the escape of M. tuberculosis from its intracellular vacuole was greatly augmented by the presence of PDIM. The increased release of M. tuberculosis into the cytosol led to increased host cell necrosis. The discovery of a link between the cell wall lipid PDIM and a major pathogenesis pathway of M. tuberculosis provides important insights into the molecular mechanisms of host cell manipulation by M. tuberculosis. IMPORTANCE Mycobacterium tuberculosis is a major human pathogen that has coevolved with its host for thousands of years. The complex and unique cell wall of M. tuberculosis contains the lipid PDIM (phthiocerol dimycocerosates), which is crucial for virulence of the bacterium, but its function is not well understood. Here we show that PDIM expression by M. tuberculosis is negatively regulated by a novel transcriptional repressor, Rv3167c. In addition, we discovered that the escape of M. tuberculosis from its intracellular vacuole was greatly augmented by the presence of PDIM. The increased release of M. tuberculosis into the cytosol led to increased host cell necrosis. The discovery of a link between the cell wall lipid PDIM and a major pathogenesis pathway of M. tuberculosis provides important insights into the molecular mechanisms of host cell manipulation by M. tuberculosis.

Expression of the Genes Encoding the Trk and Kdp Potassium Transport Systems ofMycobacterium tuberculosisduring GrowthIn Vitro

Two potassium (K+)-uptake systems, Trk and Kdp, are operative inMycobacterium tuberculosis(Mtb), but the environmental factors triggering their expression have not been determined. The current study has evaluated the expression of these genes in theMtbwild-type and atrk-gene knockout strain at various stages of logarithmic growth in relation to extracellular K+concentrations and pH. In both strains, mRNA levels of the K+-uptake encoding genes were relatively low compared to those of the housekeeping gene,sigA, at the early- and mid-log phases, increasing during late-log. Increased gene expression coincided with decreased K+uptake in the context of a drop in extracellular pH and sustained high extracellular K+concentrations. In an additional series of experiments, the pH of the growth medium was manipulated by the addition of 1N HCl/NaOH. Decreasing the pH resulted in reductions in both membrane potential and K+uptake in the setting of significant induction of genes encoding both K+transporters. These observations are consistent with induction of the genes encoding the active K+transporters ofMtbas a strategy to compensate for loss of membrane potential-driven uptake of K+at low extracellular pH. Induction of these genes may promote survival in the acidic environments of the intracellular vacuole and granuloma.

In macrophages, HIV-1 assembles into an intracellular plasma membrane domain containing the tetraspanins CD81, CD9, and CD53

In macrophages, HIV-1 has been shown to bud into intracellular structures that contain the late endosome marker CD63. We show that these organelles are not endosomes, but an internally sequestered plasma membrane domain. Using immunofluorescence microscopy and immunoelectron microscopy, we find that HIV-1 buds into a compartment that contains the tetraspanins CD81, CD9, and CD53. On uninfected macrophages, these proteins are seen at the cell surface and in intracellular vacuole-like structures with a complex content of vesicles and interconnected membranes that lack endosome markers, including CD63. Significantly, these structures are accessible to small tracers (horseradish peroxidase or ruthenium red) applied to cells at 4°C, indicating that they are connected to the cell surface. HIV assembles on, and accumulates within, these intracellular compartments. Furthermore, CD63 is recruited to the virus-containing structures and incorporated into virions. These results indicate that, in macrophages, HIV-1 exploits a previously undescribed intracellular plasma membrane domain to assemble infectious particles.

Parachlamydia acanthamoeba Enters and Multiplies within Human Macrophages and Induces Their Apoptosis

ABSTRACT Parachlamydia acanthamoeba is an obligately intracellular bacterium that naturally infects free-living amoebae. It is a potential human pathogen and may survive in human macrophages. We studied P. acanthamoeba entry into, and multiplication within, human monocyte-derived macrophages. After 8 h of incubation, 80% of macrophages were infected with a mean of 3.8 P. acanthamoeba organisms per cell. Electron microscopy demonstrated that parachlamydiae were in an intracellular vacuole. After infection with living organisms, the number of parachlamydiae per macrophage increased 4 times from day 0 to day 4, whereas heat-inactivated parachlamydiae were eliminated during the same period. Quantitative PCR confirmed that P. acanthamoeba replicates within macrophages. Transcriptional activity of P. acanthamoeba was detected by reverse transcription-PCR targeting the gene encoding ADP-ATP translocase (tlc). P. acanthamoeba exerted a cytopathic effect on macrophages. When macrophages were infected with living bacteria, their number decreased significantly from day 0 to day 4 due to apoptosis, as shown by annexin-V binding and electron microscopy. This study shows that P. acanthamoeba enters and multiplies within human macrophages before inducing their apoptosis.