Glucose Starvation Boosts Entamoeba histolytica Virulence

The unicellular parasite, Entamoeba histolytica, is exposed to numerous adverse conditions, such as nutrient deprivation, during its life cycle stages in the human host. In the present study, we examined whether the parasite virulence could be influenced by glucose starvation (GS). The migratory behaviour of the parasite and its capability to kill mammalian cells and to lyse erythrocytes is strongly enhanced following GS. In order to gain insights into the mechanism underlying the GS boosting effects on virulence, we analyzed differences in protein expression levels in control and glucose-starved trophozoites, by quantitative proteomic analysis. We observed that upstream regulatory element 3-binding protein (URE3-BP), a transcription factor that modulates E.histolytica virulence, and the lysine-rich protein 1 (KRiP1) which is induced during liver abscess development, are upregulated by GS. We also analyzed E. histolytica membrane fractions and noticed that the Gal/GalNAc lectin light subunit LgL1 is up-regulated by GS. Surprisingly, amoebapore A (Ap-A) and cysteine proteinase A5 (CP-A5), two important E. histolytica virulence factors, were strongly down-regulated by GS. While the boosting effect of GS on E. histolytica virulence was conserved in strains silenced for Ap-A and CP-A5, it was lost in LgL1 and in KRiP1 down-regulated strains. These data emphasize the unexpected role of GS in the modulation of E.histolytica virulence and the involvement of KRiP1 and Lgl1 in this phenomenon

Glucose Starvation Boosts Entamoeba histolytica

The unicellular parasite, Entamoeba histolytica, is exposed to numerous adverse conditions, such as nutrient deprivation, during its life cycle stages in the human host. In the present study, we examined whether the parasite virulence could be influenced by glucose starvation (GS). The migratory behaviour of the parasite and its capability to kill mammalian cells and to lyse erythrocytes is strongly enhanced following GS. In order to gain insights into the mechanism underlying the GS boosting effects on virulence, we analyzed differences in protein expression levels in control and glucose-starved trophozoites, by quantitative proteomic analysis. We observed that upstream regulatory element 3-binding protein (URE3-BP), a transcription factor that modulates E.histolytica virulence, and the lysine-rich protein 1 (KRiP1) which is induced during liver abscess development, are upregulated by GS. We also analyzed E. histolytica membrane fractions and noticed that the Gal/GalNAc lectin light subunit LgL1 is up-regulated by GS. Surprisingly, amoebapore A (Ap-A) and cysteine proteinase A5 (CP-A5), two important E. histolytica virulence factors, were strongly down-regulated by GS. While the boosting effect of GS on E. histolytica virulence was conserved in strains silenced for Ap-A and CP-A5, it was lost in LgL1 and in KRiP1 downregulated strains. These data emphasize the unexpected role of GS in the modulation of E.histolytica virulence and th

ATF3 Regulates the Expression of AChE During Stress

Acetylcholinesterase (AChE) expresses in non-cholinergic cells, but its role(s) there remain unknown. We have previously attributed a pro-apoptotic role for AChE in stressed retinal photoreceptors, though by unknown mechanism. Here, we examined its promoter only to find that it includes a binding sequence for the activating transcription factor 3 (ATF3); a prototypical mediator of apoptosis. This suggests that expression of AChE could be regulated by ATF3 in the retina. Indeed, ATF3 binds the AChE-promoter to down-regulate its expressions in vitro. Strikingly, retinas of “blinded” mice display hallmarks of apoptosis, almost exclusively in the outer nuclear layer (ONL); coinciding with elevated levels of AChE and absence of ATF3. A mirror image is observed in the inner nuclear layer (INL), namely prominent levels of ATF3 and lack of AChE as well as lack of apoptosis. We conclude that segregated patterns of expressions of ATF3 reflect its ability to repress apoptosis in different layers of the retina—a novel mechanism behind apoptosis

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<p>Classification of S-nitrosylated proteins in <i>Entamoeba histolytica</i> according to their biological role: Translation.</p

Proteomic Identification of <i>S</i>-Nitrosylated Proteins in the Parasite <i>Entamoeba histolytica</i> by Resin-Assisted Capture: Insights into the Regulation of the Gal/GalNAc Lectin by Nitric Oxide

<div><p><i>Entamoeba histolytica</i> is a gastrointestinal protozoan parasite that causes amebiasis, a disease which has a worldwide distribution with substantial morbidity and mortality. Nitrosative stress, which is generated by innate immune cells, is one of the various environmental challenges that <i>E. histolytica</i> encounters during its life cycle. Although the effects of nitric oxide (NO) on the regulation of gene expression in this parasite have been previously investigated, our knowledge on <i>S</i>-nitrosylated proteins in <i>E.histolytica</i> is lacking. In order to fill this knowledge gap, we performed a large-scale detection of <i>S</i>-nitrosylated (SNO) proteins in <i>E.histolytica</i> trophozoites that were treated with the NO donor, S-nitrosocysteine by resin-assisted capture (RAC). We found that proteins involved in glycolysis, gluconeogenesis, translation, protein transport, and adherence to target cells such as the heavy subunit of Gal/GalNac lectin are among the <i>S</i>-nitrosylated proteins that were enriched by SNO-RAC. We also found that the <i>S</i>-nitrosylated cysteine residues in the carbohydrate recognition domain (CRD) of Gal/GalNAc lectin impairs its function and contributes to the inhibition of <i>E.histolytica</i> adherence to host cells. Collectively, these results advance our understanding of the mechanism of reduced <i>E.histolytica</i> adherence to mammalian cells by NO and emphasize the importance of NO as a regulator of key physiological functions in <i>E.histolytica</i>.</p></div

Analysis of <i>S</i>-nitrosylated proteins in <i>E. histolytica</i> after resin-assisted capture.

<p>A. Viability of <i>E.histolytica</i> trophozoites which were exposed to different concentrations of S-nitrosocysteine (CysNO) for 20 minutes. Data are expressed as the mean and standard deviation of three independent experiments that were repeated twice. <i>E.histolytica</i> trophozoites strain HM-1:IMSS were treated with 500 µM CysNO for 20 minutes. The protein <i>S</i>-nitrosothiols (SNO) in the cell lysates was subjected to resin-assisted capture (RAC) in the presence of 40 mM ascorbate (+ASC) or the absence of ascorbate (–ASC). B. Coomassie blue staining of <i>S</i>-nitrolysated proteins. C. Functional categories of all <i>S</i>-nitrosylated proteins. <i>S</i>-nitrosylated proteins in <i>E.histolytica</i> were classified according to their biological role. D. Confirmation of <i>S</i>-nitrosylation of three proteins, enolase, glyceraldehyde-3-phosphate dehydrogenase, and the heavy subunit of Gal/GalNAc lectin after resin-assisted capture by western blotting. This figure displays a representative result from two independent experiments.</p

Classification of S-nitrosylated proteins in <i>Entamoeba histolytica</i> according to their biological role: Super-family of small GTPase.

<p>Classification of S-nitrosylated proteins in <i>Entamoeba histolytica</i> according to their biological role: Super-family of small GTPase.</p

Classification of S-nitrosylated proteins in <i>Entamoeba histolytica</i> according to their biological role: Glycolysis/Gluconeogenesis.

<p>Classification of S-nitrosylated proteins in <i>Entamoeba histolytica</i> according to their biological role: Glycolysis/Gluconeogenesis.</p