EhVps29 Has a Role in the Location of the Retromer Complex and the Function of Key Virulence Factors in Entamoeba histolytica

The retromer is a highly conserved complex that mediates the trafficking of cargo proteins to the plasma membrane or the trans-Golgi network. In pathogenic microorganisms, retromer-dependent transport contributes to the delivery of virulence factors and promotes infection. The retromer consists of a sorting nexin dimer (SNX) and a cargo-selection complex (CSC), formed by Vps26, Vps35, and Vps29. In Entamoeba histolytica, the parasite that causes human amoebiasis, the retromer functions as a Rab7A GTPase effector and participates in phagocytosis and cytotoxicity. Although we previously characterized the roles of EhVps26 and EhVps35, the function of EhVps29 remained unclear. In this study, we analyzed the subcellular localization and functional role of EhVps29 in adhesion, phagocytosis, and cytopathic effect. EhVps29 localized to the plasma membrane, cytosol, vesicles, tubules, Golgi-like structures, MVBs and, for the first time, the nucleus. Immunofluorescence and Western blot assays demonstrated that EhVps29 modulates the localization of EhVps26, EhADH adhesin, and EhCP112 cysteine protease. Ehvps29 gene silencing and overexpression confirmed its involvement in virulence-associated processes. Immunoprecipitation and confocal microscopy results showed the interaction among EhVps29 and the ESCRT machinery members EhVps36 and EhADH. Our results indicate that EhVps29 is involved in parasite virulence and protein trafficking through recycling or degradation pathways.