Biology subjects

Nakada-Tsukui, K.

Publications and source records attributed to Nakada-Tsukui, K..

3 recordsLinked to original sources

bioRxiv2026

Chromosome organization of Entamoeba histolytica and Entamoeba dispar

Entamoeba histolytica is a clinically important pathogenic eukaryote and the causative agent of amoebic dysentery. Entamoeba dispar, a nonpathogenic commensal species that resides in the human colon, is the closest sibling species, and serves as an appropriate comparator for genome-wide analysis. Although the genome of E. histolytica is approximately 26.9 Mb, and the largest known genome within the genus, that of E. invadens, is approximately 40.9 Mb, obtaining high-quality assemblies in this genus has remained challenging due to extensive repetitive regions, tRNA gene arrays, and aneuploidy. Here, we used PacBio HiFi sequencing to assemble the genomes of the pathogenic E. histolytica and the nonpathogenic E. dispar. We reconstructed all 36 chromosomes of E. histolytica and 35 chromosomes of E. dispar, assembling each as a single continuous DNA sequence (contig). The two species exhibited high genome-wide nucleotide similarity and conserved synteny at the amino acid level. At one end of each chromosome, we identified tRNA arrays, whereas the opposite end lacked such arrays, resulting in an asymmetric chromosomal architecture. Analysis of unique-read depth revealed widespread aneuploidy in both species: E. histolytica is predominantly tetraploid, whereas E. dispar is diploid, a conclusion further supported by SNP allele-frequency distributions. These assemblies provide a robust foundation for comparative genomics in Entamoeba and offer detailed insights into chromosome-end structure and ploidy.

genomics↗

bioRxiv2024

EhRacM differentially regulates macropinocytosis and motility in the enteric protozoan parasite Entamoeba histolytica

Macropinocytosis is an evolutionarily conserved endocytic process that plays a vital role in internalizing extracellular fluids and particles in cells. This non-selective endocytic pathway is crucial for various physiological functions such as nutrient uptake, sensing, signaling, antigen presentation, and cell migration. While macropinocytosis has been extensively studied in macrophages and cancer cells, the molecular mechanisms of macropinocytosis in pathogens are less understood. It has been known that Entamoeba histolytica, the causative agent of amebiasis, exploits macropinocytosis for survival and pathogenesis. Since macropinocytosis is initiated by actin polymerization, leading to the formation of membrane ruffles and the subsequent trapping of solutes in macropinosomes, actin cytoskeleton regulation is crucial. Thus, this study focuses on unraveling the role of well-conserved actin cytoskeleton regulators, Rho small GTPase family proteins, in macropinocytosis in E. histolytica. Through gene silencing of highly transcribed Ehrho/Ehrac genes and following flow cytometry analysis, we identified that silencing EhracM enhances dextran macropinocytosis and affects cellular migration persistence. Live imaging and interactome analysis unveiled the cytosolic and vesicular localization of EhRacM, along with its interaction with signaling and membrane traffic-related proteins, shedding light on EhRacMs multiple roles. Our findings provide insights into the specific regulatory mechanisms of macropinocytosis among endocytic pathways in E. histolytica, highlighting the significance of EhRacM in both macropinocytosis and cellular migration. Author SummaryEntamoeba histolytica is an intestinal protozoan parasite that causes amoebic dysentery and liver abscesses in humans. This organism exploits macropinocytosis, a cellular process that engulfs extracellular fluids and particles, for its survival and pathogenicity. Although macropinocytosis is well-characterized in immune cells and cancer cells as it is essential for nutrient uptake, its mechanisms in pathogens, such as E. histolytica, remain less explored. Our research focused on the molecular mechanisms underpinning macropinocytosis in this parasite, specifically examining the role of Rho small GTPase family proteins. These proteins are critical regulators of the actin cytoskeleton in eukaryotic cells. Our study reveals that one specific Rho small GTPase, EhRacM, is in the maturation of macropinosomes as well as in directing linear cell migration. The physiological significance of EhRacM in regulating both macropinocytosis and migration opens new avenues for understanding the role of Rho small GTPases in these signaling pathways, which could eventually lead to the development of new control measures against diseases caused by this parasite.

cell biology↗

bioRxiv2021

PTEN differentially regulates endocytosis, migration, and proliferation in the enteric protozoan parasite Entamoeba histolytica

PTEN is a lipid phosphatase that is highly conserved and involved in a broad range of biological processes including cytoskeletal reorganization, endocytosis, signal transduction, and cell migration in all eukaryotes. Although regulation of phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P3] signaling via PTEN has been well established in model organisms and mammals, it remains elusive in the parasitic protist E. histolytica, which heavily relies on PtdIns phosphate(s)-dependent membrane traffic, migration, and phago- and trogocytosis for its pathogenesis. In this study, we characterized the major PTEN from E. histolytica, EhPTEN1, which shows the highest expression at the transcript level in the trophozoite stage among 6 possible PTENs, to understand the significance of PtdIns(3,4,5)P3 signaling in this parasite. Live imaging of GFP-EhPTEN1 expressing amebic trophozoites showed localization mainly in the cytosol with a higher concentration at pseudopods and the extending edge of the phago- and trogocytic cups. Furthermore, quantitative analysis of phago- and trogocytosis using a confocal image cytometer showed that overexpression of EhPTEN1 caused reduction in trogo- and phagocytosis while transcriptional gene silencing of EhPTEN1 gene caused opposite phenotypes. These data suggest that EhPTEN1 has an inhibitory role in these biological processes. Conversely, EhPTEN1 acts as a positive regulator for fluid-phase and receptor-mediated endocytosis in E. histolytica trophozoites. Moreover, we showed that EhPTEN1 was required for optimal growth and migration of this parasite. Finally, the phosphatase activity of EhPTEN1 towards PtdIns(3,4,5)P3 was demonstrated, suggesting that the biological roles of EhPTEN1 are likely linked to its catalytic function. Taken together, these results indicate that EhPTEN1 differentially regulates multiple cellular activities essential for proliferation and pathogenesis of the organism, via PtdIns(3,4,5)P3 signaling. Elucidation of biological roles of PTEN and PtdIns(3,4,5)P3 signaling at the molecular levels promotes our understanding of the pathogenesis of this parasite and potentially leads to the design of novel therapeutics against amebiasis. Author summaryEntamoeba histolytica is an intestinal protozoan parasite that causes amoebic dysentery and liver abscesses in humans. It has been well understood how the amoebas ability to ingests and destroy human cells and invade tissues contributes to disease symptoms such as bloody diarrhea. The underlying mechanisms for such activities, called pathogenicity, include trafficking (transport) and secretion of cytolytic proteins, migration (ameboid movement), and ingestion and destruction of human cells, heavily rely on the signal transduction system via metabolism (synthesis and decomposition) of phosphoinositides (phosphatidylinositols containing 0-5 phosphates), and downstream regulation of cytoskeleton (dynamic network of interlinking protein filaments, such as actin, in the cytoplasm). In this study, we characterized one enzyme called EhPTEN1, which degrades and inactivate PtdIns(3,4,5)P3. We have shown that EhPTEN1 is involved in migration, internalization of soluble and solid materials (endocytosis, trogo-, and phagocytosis). EhPTEN1 apparently regulates cell migration, endocytosis, trogo-, phagocytosis, and proliferation in a complex fashion. Our findings help in the elucidation of the physiological significance of PTEN and cellular events regulated via phosphoinositides in this enteric parasite and other pathogenic parasites, and potentially lead to the development of new control measures against parasitic diseases.

microbiology↗