The genome and transcriptome of the enteric parasite Entamoeba invadens, a model for encystation

Abstract

BackgroundSeveral eukaryotic parasites form cysts that transmit infection. The process is found in diverse organisms such as Toxoplasma, Giardia, and nematodes. In Entamoeba histolytica this process cannot be induced in vitro, making it difficult to study. In Entamoeba invadens, stage conversion can be induced, but its utility as a model system to study developmental biology has been limited by a lack of genomic resources. We carried out genome and transcriptome sequencing of E. invadens to identify molecular processes involved in stage conversion.ResultsWe report the sequencing and assembly of the E. invadens genome and use whole transcriptome sequencing to characterize changes in gene expression during encystation and excystation. The E. invadens genome is larger than that of E. histolytica, apparently largely due to expansion of intergenic regions; overall gene number and the machinery for gene regulation are conserved between the species. Over half the genes are regulated during the switch between morphological forms and a key signaling molecule, phospholipase D, appears to regulate encystation. We provide evidence for the occurrence of meiosis during encystation, suggesting that stage conversion may play a key role in recombination between strains.ConclusionsOur analysis demonstrates that a number of core processes are common to encystation between distantly related parasites, including meiosis, lipid signaling and RNA modification. These data provide a foundation for understanding the developmental cascade in the important human pathogen E. histolytica and highlight conserved processes more widely relevant in enteric pathogens.

Regulation of gene expression in the protozoan parasite Entamoeba invadens: identification of core promoter elements and promoters with stage-specific expression patterns

“…Thus the reptilian parasite, Entamoeba invadens , for which highly efficient in vitro encystation and excystation protocols exist, has been utilized as a model system to study Entamoeba development (Vazquezdelara-Cisneros and Arroyo-Begovich, 1984; Eichinger, 1997). Using this system, it has been shown that a number of processes including lipid signaling and meiosis are developmentally regulated (Ehrenkaufer et al, 2013). Additionally, as encystation progresses, the levels of metabolites involved in glycolysis decrease drastically (De Cadiz et al, 2013).…”

“…In order to define the transcriptional network associated with stage conversion, we recently performed RNA sequencing of E. invadens during the entire developmental cycle (both encystation and excystation) (Ehrenkaufer et al, 2013). The RNA-Seq methodology was used to improve the accuracy of genome annotation and identify untranslated regions as was demonstrated in E. histolytica (Hon et al, 2013), Tetrahymena (Xiong et al, 2012) and Plasmodium (Otto et al, 2010).…”

Development of RNA Interference Trigger-Mediated Gene Silencing in Entamoeba invadens

“…Thus, the stability of silencing during the multitude of developmental changes is paramount in the use of this tool in different life cycle stages of the parasite. Recent transcriptome-sequencing (RNA-Seq) data from E. invadens has uncovered the fact that the sRNA repertoire does not change during the different developmental stages of the parasite (15). Therefore, we unfortunately did not find genes that could function as stage-specific triggers.…”

“…As the parasite undergoes a developmental shift from vegetative growth to a quiescent stage (trophozoites to cysts), approximately half of the E. invadens genome is differentially expressed at some point during the stage transition (15). Thus, it is imperative to determine if silencing is stable during this massive change in cell physiology.…”

“…An E. invadens (IP-1) culture was grown at 25°C under standard conditions (15). To establish stable transgenic lines, the parasites were transfected with 50 g plasmid by electroporation as previously described (12).…”