scholarly journals Cysteine Protease-Binding Protein Family 6 Mediates the Trafficking of Amylases to Phagosomes in the Enteric Protozoan Entamoeba histolytica

2013 ◽  
Vol 81 (5) ◽  
pp. 1820-1829 ◽  
Author(s):  
Atsushi Furukawa ◽  
Kumiko Nakada-Tsukui ◽  
Tomoyoshi Nozaki
Keyword(s):  
Entamoeba Histolytica ◽  
Cysteine Protease ◽  
Binding Protein ◽  
Proteome Analysis ◽  
Protozoan Parasite ◽  
Cysteine Proteases ◽  
Protein Family ◽  
Intestinal Protozoan ◽  
Α Subunit ◽  
Content Type

ABSTRACTPhagocytosis plays a pivotal role in nutrient acquisition and evasion from the host defense systems inEntamoeba histolytica, the intestinal protozoan parasite that causes amoebiasis. We previously reported thatE. histolyticapossesses a unique class of a hydrolase receptor family, designated the cysteine protease-binding protein family (CPBF), that is involved in trafficking of hydrolases to lysosomes and phagosomes, and we have also reported that CPBF1 and CPBF8 bind to cysteine proteases or β-hexosaminidase α-subunit and lysozymes, respectively. In this study, we showed by immunoprecipitation that CPBF6, one of the most highly expressed CPBF proteins, specifically binds to α-amylase and γ-amylase. We also found that CPBF6 is localized in lysosomes, based on immunofluorescence imaging. Immunoblot and proteome analyses of the isolated phagosomes showed that CPBF6 mediates transport of amylases to phagosomes. We also demonstrated that the carboxyl-terminal cytosolic region of CPBF6 is engaged in the regulation of the trafficking of CPBF6 to phagosomes. Our proteome analysis of phagosomes also revealed new potential phagosomal proteins.

scholarly journals The Intestinal Protozoan Parasite Entamoeba histolytica Contains 20 Cysteine Protease Genes, of Which Only a Small Subset Is Expressed during In Vitro Cultivation

2003 ◽  
Vol 2 (3) ◽  
pp. 501-509 ◽  
Author(s):  
Iris Bruchhaus ◽  
Brendan J. Loftus ◽  
Neil Hall ◽  
Egbert Tannich
Keyword(s):  
Entamoeba Histolytica ◽  
Cysteine Protease ◽  
Cathepsin L ◽  
Protozoan Parasite ◽  
Cysteine Proteases ◽  
Small Subset ◽  
In Vitro Cultivation ◽  
Intestinal Protozoan ◽  
Parasite Life Cycle

ABSTRACT Cysteine proteases are known to be important pathogenicity factors of the protozoan parasite Entamoeba histolytica. So far, a total of eight genes coding for cysteine proteases have been identified in E. histolytica, two of which are absent in the closely related nonpathogenic species E. dispar. However, present knowledge is restricted to enzymes expressed during in vitro cultivation of the parasite, which might represent only a subset of the entire repertoire. Taking advantage of the current E. histolytica genome-sequencing efforts, we analyzed databases containing more than 99% of all ameba gene sequences for the presence of cysteine protease genes. A total of 20 full-length genes was identified (including all eight genes previously reported), which show 10 to 86% sequence identity. The various genes obviously originated from two separate ancestors since they form two distinct clades. Despite cathepsin B-like substrate specificities, all of the ameba polypeptides are structurally related to cathepsin L-like enzymes. None of the previously described enzymes but 7 of the 12 newly identified proteins are unique compared to cathepsins of higher eukaryotes in that they are predicted to have transmembrane or glycosylphosphatidylinositol anchor attachment domains. Southern blot analysis revealed that orthologous sequences for all of the newly identified proteases are present in E. dispar. Interestingly, the majority of the various cysteine protease genes are not expressed in E. histolytica or E. dispar trophozoites during in vitro cultivation. Therefore, it is likely that at least some of these enzymes are required for infection of the human host and/or for completion of the parasite life cycle.

scholarly journals Localization of Phosphatidylinositol 4,5-Bisphosphate to Lipid Rafts and Uroids in the Human Protozoan Parasite Entamoeba histolytica

2013 ◽  
Vol 81 (6) ◽  
pp. 2145-2155 ◽  
Author(s):  
Amrita B. Koushik ◽  
Rhonda R. Powell ◽  
Lesly A. Temesvari
Keyword(s):  
Lipid Rafts ◽  
Entamoeba Histolytica ◽  
Protozoan Parasite ◽  
Invasive Infection ◽  
Cellular Functions ◽  
Intestinal Protozoan ◽  
Content Type ◽  
Downstream Effector ◽  
Key Factor ◽  
Actomyosin Cytoskeleton

ABSTRACTEntamoeba histolyticais an intestinal protozoan parasite and is the causative agent of amoebiasis. During invasive infection, highly motile amoebae destroy the colonic epithelium, enter the blood circulation, and disseminate to other organs such as liver, causing liver abscess. Motility is a key factor inE. histolyticapathogenesis, and this process relies on a dynamic actomyosin cytoskeleton. In other systems, phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] is known to regulate a wide variety of cellular functions, including signal transduction, actin remodeling, and cell motility. Little is known about the role of PI(4,5)P2inE. histolyticapathogenicity. In this study, we demonstrate that PI(4,5)P2is localized to cholesterol-rich microdomains, lipid rafts, and the actin-rich fractions of theE. histolyticamembrane. Microscopy revealed that the trailing edge of polarized trophozoites, uroids, are highly enriched in lipid rafts and their constituent lipid, PI(4,5)P2. Polarization and enrichment of uroids and rafts with PI(4,5)P2were enhanced upon treatment ofE. histolyticacells with cholesterol. Exposure to cholesterol also increased intracellular calcium, which is a downstream effector of PI(4,5)P2, with a concomitant increase in motility. Together, our data suggest that inE. histolytica, PI(4,5)P2may signal from lipid rafts and cholesterol may play a role in triggering PI(4,5)P2-mediated signaling to enhance the motility of this pathogen.

scholarly journals tRNA Gene Sequences Are Required for Transcriptional Silencing in Entamoeba histolytica

2009 ◽  
Vol 9 (2) ◽  
pp. 306-314 ◽  
Author(s):  
Henriette Irmer ◽  
Ina Hennings ◽  
Iris Bruchhaus ◽  
Egbert Tannich
Keyword(s):  
Entamoeba Histolytica ◽  
Trna Gene ◽  
Regulation Of Gene Expression ◽  
Transcriptional Silencing ◽  
Protozoan Parasite ◽  
Upstream Region ◽  
Trna Genes ◽  
Intestinal Protozoan ◽  
Content Type ◽  
Systematic Assessment

ABSTRACT Transcriptional silencing by trans inactivation can contribute to the regulation of gene expression in eukaryotic cells. In the human intestinal protozoan parasite Entamoeba histolytica, trans inactivation of the amoebapore-A gene (AP-A) was recently achieved by episomal transfection of E. histolytica trophozoites with the plasmid psAP1. The mechanism of AP-A trans inactivation is largely unknown, though it was suggested that a partial short interspersed transposable element (SINE) is required. By systematic assessment of various E. histolytica isolates transfected with psAP1 derivates, trans inactivation of AP-A was restricted to the strain HM-1:IMSS (2411) but could not be achieved in other standard laboratory strains. Importantly, sequences of an E. histolytica tRNA array that were located on psAP1 in close proximity to the AP-A upstream region and comprising the glutamic acid (TTC) (E) and tyrosine (GTA) (Y) tRNA genes were indispensable for AP-A silencing. In contrast to the case described in previous reports, SINE was not required for AP-A trans inactivation. AP-A expression could be regained in silenced cells by episomal transfection under the control of a heterologous E. histolytica promoter, opening a way toward future silencing of individual genes of interest in E. histolytica. Our results indicate that tRNA gene-mediated silencing is not restricted to Saccharomyces cerevisiae.

scholarly journals Localization and Targeting of an Unusual Pyridine Nucleotide Transhydrogenase in Entamoeba histolytica

2010 ◽  
Vol 9 (6) ◽  
pp. 926-933 ◽  
Author(s):  
Mohammad Abu Yousuf ◽  
Fumika Mi-ichi ◽  
Kumiko Nakada-Tsukui ◽  
Tomoyoshi Nozaki
Keyword(s):  
Entamoeba Histolytica ◽  
Fluorescent Protein ◽  
Physiological Role ◽  
Protozoan Parasite ◽  
Pyridine Nucleotide ◽  
Immunofluorescence Assay ◽  
Chemical Degradation ◽  
Content Type ◽  
Targeting Signal ◽  
Pyridine Nucleotide Transhydrogenase

ABSTRACT Pyridine nucleotide transhydrogenase (PNT) catalyzes the direct transfer of a hydride-ion equivalent between NAD(H) and NADP(H) in bacteria and the mitochondria of eukaryotes. PNT was previously postulated to be localized to the highly divergent mitochondrion-related organelle, the mitosome, in the anaerobic/microaerophilic protozoan parasite Entamoeba histolytica based on the potential mitochondrion-targeting signal. However, our previous proteomic study of isolated phagosomes suggested that PNT is localized to organelles other than mitosomes. An immunofluorescence assay using anti-E. histolytica PNT (EhPNT) antibody raised against the NADH-binding domain showed a distribution to the membrane of numerous vesicles/vacuoles, including lysosomes and phagosomes. The domain(s) required for the trafficking of PNT to vesicles/vacuoles was examined by using amoeba transformants expressing a series of carboxyl-terminally truncated PNTs fused with green fluorescent protein or a hemagglutinin tag. All truncated PNTs failed to reach vesicles/vacuoles and were retained in the endoplasmic reticulum. These data indicate that the putative targeting signal is not sufficient for the trafficking of PNT to the vesicular/vacuolar compartments and that full-length PNT is necessary for correct transport. PNT displayed a smear of >120 kDa on SDS-PAGE gels. PNGase F and tunicamycin treatment, chemical degradation of carbohydrates, and heat treatment of PNT suggested that the apparent aberrant mobility of PNT is likely attributable to its hydrophobic nature. PNT that is compartmentalized to the acidic compartments is unprecedented in eukaryotes and may possess a unique physiological role in E. histolytica.

scholarly journals Autophagy Activated by Peroxiredoxin of Entamoeba histolytica

Cells ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 2462
Author(s):  
Xia Li ◽  
Yuhan Zhang ◽  
Yanqing Zhao ◽  
Ke Qiao ◽  
Meng Feng ◽  
...  
Keyword(s):  
Entamoeba Histolytica ◽  
Protozoan Parasite ◽  
Host Cells ◽  
Functional Domain ◽  
Intestinal Protozoan ◽  
Raw264.7 Macrophages ◽  
Evolutionarily Conserved ◽  
Amoebic Colitis ◽  
Dependent Cell ◽  
Cellular Components

Autophagy, an evolutionarily conserved mechanism to remove redundant or dangerous cellular components, plays an important role in innate immunity and defense against pathogens, which, in turn, can regulate autophagy to establish infection within a host. However, for Entamoeba histolytica, an intestinal protozoan parasite causing human amoebic colitis, the interaction with the host cell autophagy mechanism has not been investigated. In this study, we found that E. histolytica peroxiredoxin (Prx), an antioxidant enzyme critical for parasite survival during the invasion of host tissues, could activate autophagy in macrophages. The formation of autophagosomes in macrophages treated with recombinant Prx of E. histolytica for 24 h was revealed by immunofluorescence and immunoblotting in RAW264.7 cells and in mice. Prx was cytotoxic for RAW264.7 macrophages after 48-h treatment, which was partly attributed to autophagy-dependent cell death. RNA interference experiments revealed that Prx induced autophagy mostly through the toll-like receptor 4 (TLR4)–TIR domain-containing adaptor-inducing interferon (TRIF) pathway. The C-terminal part of Prx comprising 100 amino acids was the key functional domain to activate autophagy. These results indicated that Prx of E. histolytica could induce autophagy and cytotoxic effects in macrophages, revealing a new pathogenic mechanism activated by E. histolytica in host cells.

scholarly journals Near-chromosome level genome assembly reveals ploidy diversity and plasticity in the intestinal protozoan parasite Entamoeba histolytica

BMC Genomics ◽  
2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Tetsuro Kawano-Sugaya ◽  
Shinji Izumiyama ◽  
Yasuaki Yanagawa ◽  
Yumiko Saito-Nakano ◽  
Koji Watanabe ◽  
...  
Keyword(s):  
Entamoeba Histolytica ◽  
Evolutionary Biology ◽  
Regulation Of Gene Expression ◽  
Protozoan Parasite ◽  
Model Organisms ◽  
Intestinal Protozoan ◽  
Disease Symptoms ◽  
Veterinary Importance ◽  
Reference Genomes ◽  
Chromosome Level

Abstract Background Amoebozoa is a eukaryotic supergroup composed of unicellular and multicellular amoebic protozoa (e.g. Acanthamoeba, Dictyostelium, and Entamoeba). They are model organisms for studies in cellular and evolutionary biology and are of medical and veterinary importance. Despite their importance, Amoebozoan genome organization and genetic diversity remain poorly studied due to a lack of high-quality reference genomes. The slime mold Dictyostelium discoideum is the only Amoebozoan species whose genome is available at the chromosome-level. Results Here, we provide a near-chromosome-level assembly of the Entamoeba histolytica genome, the second semi-completed Amoebozoan genome. The availability of this improved genome allowed us to discover inter-strain heterogeneity in ploidy at the near-chromosome or sub-chromosome level among 11 clinical isolates and the reference strain. Furthermore, we observed ploidy-independent regulation of gene expression, contrary to what is observed in other organisms, where RNA levels are affected by ploidy. Conclusions Our findings offer new insights into Entamoeba chromosome organization, ploidy, transcriptional regulation, and inter-strain variation, which will help to further decipher observed spectrums of virulence, disease symptoms, and drug sensitivity of E. histolytica isolates.

scholarly journals Conserved Pyridoxal 5'-Phosphate-Binding Protein YggS Impacts Amino Acid Metabolism through Pyridoxine 5'-Phosphate inEscherichia coli

2019 ◽  
Vol 85 (11) ◽  
Author(s):  
Tomokazu Ito ◽  
Kana Yamamoto ◽  
Ran Hori ◽  
Ayako Yamauchi ◽  
Diana M. Downs ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Biosynthetic Pathway ◽  
Acid Metabolism ◽  
Binding Protein ◽  
Protein Family ◽  
Pleiotropic Effects ◽  
Vitamin B ◽  
Content Type ◽  
E Coli

ABSTRACTEscherichia coliYggS (COG0325) is a member of the highly conserved pyridoxal 5′-phosphate (PLP)-binding protein (PLPBP) family. Recent studies suggested a role for this protein family in the homeostasis of vitamin B6and amino acids. The deletion or mutation of a member of this protein family causes pleiotropic effects in many organisms and is causative of vitamin B6-dependent epilepsy in humans. To date, little has been known about the mechanism by which lack of YggS results in these diverse phenotypes. In this study, we determined that the pyridoxine (PN) sensitivity observed inyggS-deficientE. coliwas caused by the pyridoxine 5′-phosphate (PNP)-dependent overproduction of Val, which is toxic toE. coli. The data suggest that theyggSmutation impacts Val accumulation by perturbing the biosynthetic of Thr from homoserine (Hse). Exogenous Hse inhibited the growth of theyggSmutant, caused further accumulation of PNP, and increased the levels of some intermediates in the Thr-Ile-Val metabolic pathways. Blocking the Thr biosynthetic pathway or decreasing the intracellular PNP levels abolished the perturbations of amino acid metabolism caused by the exogenous PN and Hse. Our data showed that a high concentration of intracellular PNP is the root cause of at least some of the pleiotropic phenotypes described for ayggSmutant ofE. coli.IMPORTANCERecent studies showed that deletion or mutation of members of the YggS protein family causes pleiotropic effects in many organisms. Little is known about the causes, mechanisms, and consequences of these diverse phenotypes. It was previously shown thatyggSmutations inE. coliresult in the accumulation of PNP and some metabolites in the Ile/Val biosynthetic pathway. This work revealed that some exogenous stresses increase the aberrant accumulation of PNP in theyggSmutant. In addition, the current report provides evidence indicating that some, but not all, of the phenotypes of theyggSmutant inE. coliare due to the elevated PNP level. These results will contribute to continuing efforts to determine the molecular functions of the members of the YggS protein family.

Sign in / Sign up

Export Citation Format

Share Document