Transcriptional profiling of Entamoeba histolytica trophozoites

MacFarlane, Ryan C.; Shah, Preetam H.; Singh, Upinder

doi:10.1016/j.ijpara.2005.02.006

Cited by 32 publications

(27 citation statements)

References 67 publications

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“…Amebae were harvested, and total RNA was extracted using TRIzol reagent (Invitrogen) and cleaned using the RNeasy cleanup kit (QIAGEN). Northern blot tests were performed using standard protocols (37). Briefly, 10 to 20 g of total parasite RNA was separated by denaturing 1.2% agarose gel electrophoresis, transferred to membrane filters, and hybridized with radioactive probes using the ExpressHyb (Clontech, Palo Alto, CA) hybridization buffer.…”

Section: Methodsmentioning

confidence: 99%

“…Whether these genomic differences contribute to the various virulence phenotypes remains to be determined. Previous studies have shown that E. histolytica HM-1:IMSS is able to lyse colonic cell monolayers without major changes in its transcriptional profile, indicating that many of the genes important in host cell damage may be constitutively expressed under tissue culture conditions (37).…”

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confidence: 99%

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Identification of Differentially Expressed Genes in Virulent and NonvirulentEntamoebaSpecies: Potential Implications for Amebic Pathogenesis

2006

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Entamoeba histolytica is a protozoan parasite that causes colitis and liver abscesses. Several Entamoeba species and strains with differing levels of virulence have been identified. E. histolytica HM-1:IMSS is a virulent strain, E. histolytica Rahman is a nonvirulent strain, and Entamoeba dispar is a nonvirulent species. We used an E. histolytica DNA microarray consisting of 2,110 genes to assess the transcriptional differences between these species/strains with the goal of identifying genes whose expression correlated with a virulence phenotype. We found 415 genes expressed at lower levels in E. dispar and 32 genes with lower expression in E. histolytica Rahman than in E. histolytica HM-1:IMSS. Overall, 29 genes had decreased expression in both the nonvirulent species/strains than the virulent E. histolytica HM-1:IMSS. Interestingly, a number of genes with potential roles in stress response and virulence had decreased expression in either one or both nonvirulent Entamoeba species/strains. These included genes encoding Fe hydrogenase (9.m00419), peroxiredoxin (176.m00112), type A flavoprotein (6.m00467), lysozyme (6.m00454), sphingomyelinase C (29.m00231), and a hypothetical protein with homology to both a Plasmodium sporozoite threonine-asparagine-rich protein (STARP) and a streptococcal hemagglutinin (238.m00054). The function of these genes in Entamoeba and their specific roles in parasite virulence need to be determined. We also found that a number of the non-long-terminal-repeat retrotransposons (EhLINEs and EhSINEs), which have been shown to modulate gene expression and genomic evolution, had lower expression in the nonvirulent species/strains than in E. histolytica HM-1:IMSS. Our results, identifying expression profiles and patterns indicative of a virulence phenotype, may be useful in characterizing the transcriptional framework of virulence.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Identification of Differentially Expressed Genes in Virulent and NonvirulentEntamoebaSpecies: Potential Implications for Amebic Pathogenesis

2006

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“…Northern blots were performed as previously described (32). Briefly, total parasite RNA was isolated using TRIZOL reagent (Invitrogen) according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

“…RNA (10 to 20 g) was separated by denaturing 1.2% agarose gel electrophoresis, transferred to Hybond-N membrane filters (Amersham-Pharmacia), and hybridized with radioactive probes using the ExpressHyb (Clontech) hybridization buffer. PCR product probes were sequenced and labeled with [␣- 32 P]dATP with a random primed DNA labeling kit (Roche). Oligonucleotide probes were labeled with [␥-32 P]dATP and T4 polynucleotide kinase (New England Biolabs).…”

Section: Methodsmentioning

confidence: 99%

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Identification of an Entamoeba histolytica Serine-, Threonine-, and Isoleucine-Rich Protein with Roles in Adhesion and Cytotoxicity

MacFarlane

Singh

2007

Eukaryot Cell

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Entamoeba histolytica is a leading cause of parasitic death globally. However, the molecular framework regulating pathogenesis is poorly understood. We have previously used expression profiling to identify Entamoeba genes whose expressions were strictly associated with virulent strains (R. C. MacFarlane and U. Singh, Infect. Immun. 74:340-351, 2006). One gene, which we have named EhSTIRP (Entamoeba histolytica serine-, threonine-, and isoleucine-rich protein), was exclusively expressed in virulent but not in nonvirulent Entamoeba strains. EhSTIRP is predicted to be a transmembrane protein and is encoded by a multigene family. In order to characterize its function in amebic biology, we used a double-stranded RNA-based approach and were able to selectively down-regulate expression of this gene family. Upon EhSTIRP down-regulation, we were able to ascribe cytotoxic and adhesive properties to the protein family using lactate dehydrogenase release and Chinese hamster ovary cell adhesion assays. EhSTIRP thus likely represents a novel determinant of virulence in Entamoeba histolytica. This work validates the fact that genes expressed exclusively in virulent strains may represent virulence determinants and highlights the need for further functional analyses of other genes with similar expression profiles.Entamoeba histolytica is a protozoan parasite that causes significant morbidity and mortality in the developing world (48). The life cycle is composed of two stages: the transmissible cyst that is released in the feces of infected humans, and the motile trophozoite, which resides in the large bowel. The infection begins when cysts are ingested, usually through contaminated food or water. Excystation occurs in the small bowel, and the motile trophozoites eventually colonize the large bowel.The majority of individuals with E. histolytica infection remain asymptomatic; however, in some cases the parasites penetrate the colon wall causing dysentery and colitis and occasionally disseminate hematogenously to cause liver abscesses. The potential reasons for why so many individuals remain asymptomatic include differences in host immunity and gut microflora as well as genetic differences between Entamoeba isolates. The molecular processes that underlie the progression from gut commensal to pathogen are not completely understood (46). Invasion begins when the parasite attaches to host cells via the galactose/N-acetylgalactosamine (Gal/GalNAc)-inhibitable lectin. This binding event is followed by rapid cell death, which has been characterized as having both necrotic and apoptotic features (9, 44). Additional molecules important in amebic pathogenesis include the cysteine proteinases, papain family proteases that have been proven to be important in virulence. Parasites genetically engineered to produce low levels of cysteine proteinase activity showed significantly less gut inflammation and were unable to form liver abscesses in animal models of amebic disease (4, 5, 51). The amoebapores, pore-forming peptides, insert into the memb...

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Current awareness on comparative and functional genomics

2005

Comp Funct Genom

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In order to keep subscribers up‐to‐date with the latest developments in their field, this current awareness service is provided by John Wiley & Sons and contains newly‐published material on comparative and functional genomics. Each bibliography is divided into 16 sections. 1 Reviews & symposia; 2 General; 3 Large‐scale sequencing and mapping; 4 Genome evolution; 5 Comparative genomics; 6 Gene families and regulons; 7 Pharmacogenomics; 8 Large‐scale mutagenesis programmes; 9 Functional complementation; 10 Transcriptomics; 11 Proteomics; 12 Protein structural genomics; 13 Metabolomics; 14 Genomic approaches to development; 15 Technological advances; 16 Bioinformatics. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted

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Transcriptional profiling of Entamoeba histolytica trophozoites

Cited by 32 publications

References 67 publications

Identification of Differentially Expressed Genes in Virulent and NonvirulentEntamoebaSpecies: Potential Implications for Amebic Pathogenesis

Identification of Differentially Expressed Genes in Virulent and NonvirulentEntamoebaSpecies: Potential Implications for Amebic Pathogenesis

Identification of an Entamoeba histolytica Serine-, Threonine-, and Isoleucine-Rich Protein with Roles in Adhesion and Cytotoxicity

Current awareness on comparative and functional genomics

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