EhVps23, an ESCRT-I Member, Is a Key Factor in Secretion, Motility, Phagocytosis and Tissue Invasion by Entamoeba histolytica

Abstract

The EhVps23 protein, an orthologue of the yeast Vps23 and the mammalian TSG101 proteins, is the single member of the ESCRT-I complex of Entamoeba histolytica identified and characterized until now. EhVps23 actively participates in vesicular trafficking and phagocytosis, which influence several cellular events. In this paper, we investigated the role of EhVps23 in virulence-related functions, including the invasive capacity of trophozoites, using transfected trophozoites. Trophozoites overexpressing the EhVps23 protein (Neo-EhVps23) presented helical arrangements in the cytoplasm, similar to the ones formed by EhVps32 for scission of vesicles. By confocal and transmission electron microscopy, EhVps23 was detected in multivesicular bodies, vesicles, and the extracellular space. It was secreted in vesicles together with other proteins, including the EhADH adhesin. Probably, these vesicles carry molecules that participate in the prey capture or in cell-cell communication. Mass spectrometry of precipitates obtained using α-EhVps23 antibodies, evidenced the presence of proteins involved in motility, phagocytosis, vesicular trafficking and secretion. The study of cellular functions, revealed that Neo-EhVps23 trophozoites exhibit characteristics similar to those described for mammalian transformed cells: they grew 50% faster than the control; presented a significant higher rate of phagocytosis, and migrated five-fold faster than the control, in concordance with the low rate of migration exhibited by Ehvps23-knocked down trophozoites. In addition, Neo-EhVps23 trophozoites produced dramatic liver abscesses in experimental animals. In conclusion, our results showed that EhVps23 overexpression gave to the trophozoites characteristics that resemble cancer cells, such as increased cell proliferation, migration, and invasion. The mutant that overexpresses EhVps23 can be a good study model to explore different events related to the transformation of malignant cells.

Molecular interplays of the Entamoeba histolytica endosomal sorting complexes required for transport during phagocytosis

“…As EhADH, EhVps23 could play a role at various points of the endocytic pathway. Another experimental data on the relevance of EhVps23 in phagocytosis is the fact that trophozoites overexpressing EhVps23 migrate five-fold faster than control parasites, in concordance with the low rate of migration exhibited by Ehvps23-knocked down trophozoites (Galindo et al, 2022); this, points out to the participation of the ESCRT machinery also in motility, a fundamental event for phagocytosis. In regard to EhVps37, Loṕez-Reyes et al ( 2010) found an EhVps37D protein lacking the alpha helixes required for ESCRT-I binding (Kostelansky et al, 2007;Galindo et al, 2021).…”

“…and mass spectrometry analysis, our group detected EhVps26 and EhVps35 as interacting partners of EhVps23 (Galindo et al, 2022).…”

“…The ESCRT proteins interact with other molecules and together, they construct a chain of events that maintain the continuity of the process. Each one of the events and molecules of this chain are important, and if one of them is affected, the entire process is disturbed ( López-Reyes et al., 2011 ; Avalos-Padilla et al., 2018 ; Galindo et al., 2022 ).…”

“…ESCRT proteins change their cell location through phagocytosis, according to the advance of the event ( López-Reyes et al., 2010 ). Their knock down or overexpression have repercussions in the rate of phagocytosis and in the in vitro and in vivo virulence expression ( López-Reyes et al., 2010 ; Bañuelos et al., 2012 ; Avalos-Padilla et al., 2015 ; Avalos-Padilla et al., 2018 ; Galindo et al., 2021 ; Galindo et al., 2022 ), highlighting their role in membrane transformation and in vesicles, tubes and tunnel-like structures formation, which in turn contribute to the transport of cargo, or the prey.…”

“…In general, Rab small GTPases play a fundamental role in signalling and activation of distinct molecules involved in the process ( Saito-Nakano et al., 2005 ; Verma et al., 2020 ). ESCRT proteins have also been detected in phagosomes, late endosomes (LE), multivesicular bodies (MVBs) and their intraluminal vesicles (ILVs), in secreted vesicles, and in the MVBs fused to lysosomes ( López-Reyes et al., 2010 ; Bañuelos et al., 2012 ; Avalos-Padilla et al., 2015 ; Avalos-Padilla et al., 2018 ; Galindo et al., 2021 ; Galindo et al., 2022 ). These sequential events allow first, the transport of the prey through compartments containing different enzymes and molecules, ending in the prey digestion for recycling useful proteins, or in the secretion of others.…”