Entamoeba histolytica causes amoebiasis and damages intestinal epithelium, but how individual parasite factors coordinate parasite gene regulation with host remodeling is unclear. We investigate lysine- and glutamic acid-rich protein 2, a factor linked to the host brush border. Sequence analysis, imaging, and functional assays show that lysine- and glutamic acid-rich protein 2 accumulates in the parasite nucleus, binds AT-rich DNA, and modulates transcriptional programs related to Amoebiasis, including cysteine protease and sulfur metabolism. During parasite-epithelium contact, the protein enters host epithelial cells and is associated with increased deoxyribonucleic acid synthesis, altered cytoskeletal regulators, actin remodeling, and reduced barrier integrity. Here, we propose a working model in which lysine- and glutamic acid-rich protein 2 links parasite chromatin-associated regulation with host epithelial remodeling during contact. Notably, our data support host cytoskeletal and junctional phenotypes and do not yet establish a direct role for it in host chromatin regulation. Together, these observations suggest a potentially broader mechanism by which extracellular pathogens deploy effectors to optimize virulence and adapt to diverse host environments. © 2026. The Author(s).

Tetraspanins (TSPANs) are a family of proteins highly conserved in all eukaryotes. Although protein-protein interactions of TSPANs have been well established in eukaryotes including parasitic protists, the role they play in parasitism and pathogenesis remains largely unknown. In this study, we characterized three representative members of TSPANs, TSPAN4, TSPAN12, and TSPAN13 from the human intestinal protozoan Entamoeba histolytica. Co-immunoprecipitation assays demonstrated that TSPAN4, TSPAN12 and TSPAN13 are reciprocally pulled down together with several other TSPAN-interacting proteins including TSPAN binding protein of 55kDa (TBP55) and interaptin. Blue native-PAGE analysis showed that these TSPANs form several complexes of 120-250 kDa. Repression of tspan12 and tspan13 gene expression led to decreased secretion of cysteine proteases, while repression of tspan4 led to a four-fold increase in the activity of cysteine proteases in crude extracellular vesicles (EVs) fraction. Meanwhile, strains overexpressing HA-tagged TSPAN12 and TSPAN13 demonstrated reduced adhesion to collagen. Altogether, this study reveals that the TSPANs, especially TSPAN12 and TSPAN13, are engaged with complex protein-protein interactions and are involved in the pathogenicity-related biological functions such as protease secretion and adhesion, offering insights into the potential regulatory mechanisms of tetraspanins in protozoan parasites.