Adaptation of E. histolytica to long term glucose starvation
Ontology highlight
ABSTRACT:
ORGANISM(S): Entamoeba histolytica
SUBMITTER: Christian Weber
PROVIDER: E-MTAB-1187 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:The ability of Entamoeba histolytica to phagocytose host cells correlates to observed virulence in vivo. To better understand the mechanism of phagocytosis we used paramagnetic beads coated with host ligand and sorted trophozoites based on phagocytic ability. Gene expression was then measured in both the sorted phagocytic and non-phagocytic populations using a custom Affymetrix chip for E. histolytica. Feed forward regulation of phagocytosis by Entamoeba histolytica. Infection and Immunity. PMID 23045476 M280 Streptavidin Dynabeads (Invitrogen) were labeled with 20ug/mL biotinylated Human C1q (Quidel). Entamoeba histolytica (strain HM1) were washed twice with PBS then resuspended with the labeled beads at a 10:1 ratio of beads to trophozoites. Samples were incubated at 37°C for 45 minutes, washed twice with agitation to remove adherent beads, then resuspended in MACS buffer. Samples were loaded into magnetic columns (Miltenyi Biotec) and trophozoites were seperated according to manufacturer's protocols. phagocytic vs. non-phagocytic Entamoeba histolytica populations
2012-10-16 | E-GEOD-41604 | biostudies-arrayexpress
Project description:Entamoeba histolytica contains a large and novel family of transmembrane kinases (TMK) with proposed roles that include both amebic response to the environment and immune evasion. In the later case, the process requires several elements --these include a large gene family encoding antigenically distinct surface proteins and the expression of one variant antigen at a time by a single pathogen. Laser-capture microdissection was used in conjunction with microarray analysis to demonstrate that single trophozoites express more than one TMK gene. Overall, our data indicates that multiple members of the novel E. histolytica TMK family are utilized for non-redundant functions by the parasite. For laser capture microdissection analysis, harvested trophozoites were allowed to adhere to metal framed PEN membrane slides (Molecular Devices, Sunnyvale, CA) for 15 mins at 37°C in TYI-S-33 media. Subsequently, single cells were captured from the PEN slides using a Pix Cell II laser capture microdissection system equipped with an Olympus microscope (Arcturus Engineering, Mountain View, CA).
2010-11-01 | E-GEOD-19064 | biostudies-arrayexpress
Project description:To investigate the magnitude of the transcriptional changes occurring during the life cycle of Giardia lamblia we compared the transcriptome of trophozoites and cysts. Cysts were found to possess a much smaller transcriptome, both in terms of mRNA diversity and abundance. Genes encoding proteins related to ribosomal functions are highly over-represented. In cysts which have lost infectivity the transcriptome is further depleted. In contrast, exposure of cysts to conditions which promote excystation induced transcription. Six cysts and three trophozoites life cycle stages were analyzed.
2011-11-09 | E-GEOD-25460 | biostudies-arrayexpress
Project description:We previously showed that epigenetic transcriptional silencing of the amoebapore gene (Ehap-a) in E. histolytica G3 trophozoites was accompanied by the down-regulation of two other members of the saposin-like gene family (Ehap-b, and EhSaplip1). Comparison of the transcriptomes of G3 trophozoites with those of the parent HM-1:IMSS strain revealed 72 additional transcripts that were either down- or up-regulated at least 7 fold. One of the polyA+, up-regulated (>200) transcripts (459.m00030), had the potential to encode a 66 amino acid protein. Antibodies raised against a synthetic N-terminal peptide of the gene did not recognize any such protein in trophozoite lysates. Trophozoite cultures grown with the proteasome inhibitor MG-132 to prevent protein degradation, also did not have any detectable protein. Transfections with a plasmid in which the Ehactin gene promoter elements were used to flank the 459.m gene, significantly increased their level transcript but again, no protein was detected. Transfectants in which the 459.m ORF was placed downstream to the GST gene, expressed a fused protein which reacted with the antibody against the m.459 peptide. Constructs with the CAT reporter gene placed under the 459.m gene promoter element or fused to the C-terminal sequence of the 459.m ORF were prepared to determine if the higher levels of 459.m transcript found in the G3 trophozoites were due to changes in the stability of the transcript or increased transcription. No difference was observed in the very weak expressions seen in both G3 and HM-1:IMSS transfectants. Similarly, transfectants with a plasmid construct in which the 459.m gene was placed under its regulatory elements, did not induce an increase in the transcript levels. Our conclusions are that the transcript of the 459.m gene is an mRNA-like ncRNA and that modulations of transcript levels, especially of hypothetical genes, may not correlate with changes in the proteome. Samples include two replicates of the HM-1:IMSS strain and the amoebapore a silenced G3 strain (G3(A))
2011-08-30 | E-GEOD-28883 | biostudies-arrayexpress
Project description:Giardia causes more episodes of illness worldwide than any other parasite. It is a flagellated cyst-forming enteric pathogen that inhabits the lumen of the small intestine and although it is clear that more and less virulent strains do occur, pathogenesis and virulence of Giardia remains poorly understood. Identification of Giardia virulence factors is highly desirable since their expression would be the best indicator for determining disease outcome. The advent of quantitative proteomics has been timely in investigating which proteins are likely to be secreted virulence factors. Here we successfully apply such analyses to the whole parasite and to the supernatants derived from the parasite, in order to ascertain a repertoire of secreted proteins. In our initial analysis we compared replicates of Giardia cells and culture supernatants from the two major lineages which cause human disease – assemblage A and assemblage B. The genome of Giardia is believed to contain open reading frames which could encode as many as 6000 proteins. In our study we confirm expression of ~1600 proteins from each assemblage, the vast majority of which being common to both lineages. To look for actual enrichment of secreted proteins, we considered the ratio of proteins in the supernatant compared with the pellet. This simple method defines a small group of enriched proteins, putatively secreted at a steady state by cultured growing Giardia trophozoites of both assemblages. This secretome contains a high proportion annotated to have N’ terminus signal peptides, with some showing evidence of having recently evolved under strong positive selective pressure. The most abundant secreted proteins include known virulence factors such as the Cathepsin B cysteine proteases and members of a Giardia superfamily of Cysteine Rich Proteins which comprises VSPs, HCMPs and a new class of virulence factors, the Giardia Tenascins. Since we also find that physiological function of human enteric epithelial cells can be disrupted by soluble factors even in the absence of the trophozoites we are able to propose a straightforward model of Giardia pathogenesis incorporating key roles for the major Giardia derived soluble mediators.
2018-01-10 | PXD004398 | Pride
Project description:Evaluation of transcript levels of two strains E.Histolytica (virulent and avirulent) in two conditions (in culture or in contact with a human colon explant)
2012-08-01 | E-MTAB-1201 | biostudies-arrayexpress
Project description:Entamoeba histolytica is a human pathogen, responsible for amoebic dysentery and invasive extraintestinal disease. The parasite faces several types of stress within the host, and to establish a successful infection it must generate a robust adaptive response against host defense mechanisms. In order to obtain comprehensive information of gene expression changes in E. histolytica under growth stress, we have (1) optimized the proteomics protocol to increase the protein coverage in E. histolytica, and (2) integrated proteomic data with transcriptomic analysis under the same conditions. We have applied this approach to better understand the cellular response during serum-starvation. Label-free quantitative proteomics was performed, and compared with mRNA levels based on RNA-seq data to decipher regulation at translational and transcriptional levels. Across all samples, 2344 proteins were identified, which is an improvement over the maximum recorded number in E. histolytica proteomic studies so far. A total of 127 proteins were found to be differentially expressed and associated with functions including antioxidant activity, cytoskeleton, translation, catalysis, and transport, which revealed proteomic signatures to distinguish serum-starved from normal trophozoites. Gal/GalNAc-inhibitable lectin, Lgls, Hgl3 and Igl were repeatedly identified as significantly altered in serum-stress condition. Further, integration of transcriptomic and proteomic data revealed instances of post-transcriptional regulation. Six highly expressed transcripts had low corresponding protein expression, indicating translational repression. Conversely, eleven transcripts showed much greater downregulation compared with their corresponding proteins, indicating translational induction, or increased stability of these proteins during serum stress in E. histolytica. This multi-omics approach enables more refined gene expression analysis that would not be possible at the mRNA or protein levels alone. Our study provides important data to further understand the adaptive response of E. histolytica to growth stress.
2022-09-23 | PXD031921 | Pride
Project description:Amongst several amoebic species that colonize the human gut only Entamoeba histolytica is able to cause tissue damage in the intestine and other organs. Amoebic pathogenicity and virulence are commonly evaluated as the parasiteM-+s ability to produce liver abscesses in hamsters (ALAH). Some molecules involved in several functions that enable E. histolytica to invade and survive in the tissues have been identified and proposed as virulence factors. However, despite the fact that during early tissue invasion E. histolytica has to cope with toxic oxygen concentrations, little attention have received the mechanisms of oxygen resistance and its relationship with pathogenicity and virulence. In this work we compared the ability of virulent E. histolytica, non virulent E. histolytica and non pathogenic E. dispar to survive under physiological oxygen concentrations. Also, the effect of hyperoxia on the transcriptome, the oxygen reduction pathway, iron-sulfur protein oxidation, protein carbonylation and complement resistance was analyzed. Our results showed that a low oxygen reduction capacity plus high complement susceptibility contribute to the inability of E. dispar to survive during the early tissue invasion. On the other hand, under hyperoxia, the non virulent E. histolytica phenotype was associated with a defect in the up-regulation of genes of the heat-shock response which correlated with accumulation of oxidized proteins and irreversible PFOR inactivation. We conclude that maintenance of an intracellular hypoxic environment and up regulation of the heat-shock protein response for proper metabolic functioning are primary requirements for amoebic pathogenicity and virulence.
2015-03-02 | E-MTAB-1207 | biostudies-arrayexpress
Project description:Amoebiasis is a disease caused by the protozoan parasite, Entamoeba histolytica, with or without clinical symptoms . It has been suggested that pathogenic amoebae use three major virulence factors: Gal/GalNAc lectin, cysteine proteinases and amoebapores. However, these molecules cannot exclusively be responsible for amoebic virulence, because most of them are found in pathogenic E. histolytica as well as in non-pathogenic E. dispar, a not close correlation has been found in amoebic strains with different virulence phenotype. Due to the high genetic variability of E. histolytica isolates and strains, differential gene expression might simply reflect inter-isolate genetic variation rather than specific differences linked to virulence. In the current study, we obtained a non-virulent UG10 mutant derived from E. histolytica HM-1:IMSS. Comparing the transcriptome of both strains, no differences in gene expression of the classical virulence factors were observed. RAB family GTPase and AIG1 family members showed higher transcriptional levels in the virulent strain than in the non-virulent mutant. Our study suggests a number of novel gene products in E. histolytica with a role yet to determine in cell physiology and involvement in the pathogenesis process.
2015-05-15 | E-MTAB-3525 | biostudies-arrayexpress
Project description:Entamoeba histolytica is the parasite causing amoebiasis, an infectious disease targeting the intestine and liver of humans. The molecules involved in amoeba adaptation to different iron conditions during the invasive process are unknown. To investigate the effects of iron availability on gene expression in E. histolytica, we determined by microarray experiments the gene expression profile of parasites grown in the presence of different iron concentrations. Conditions included low amounts of iron, iron starvation and iron starvation supplemented with hemoglobin (Hb). Genes encoding important proteins involved in iron metabolism, reported in other organism such as bacteria, were identified for the first time in E. histolytica. The influence of iron on the amount of these transcripts was confirmed by quantitative real-time PCR and their protein products were analyzed by Western blots
2014-06-30 | E-MTAB-1158 | biostudies-arrayexpress