Proteomic Study of Entamoeba histolytica Trophozoites, Cysts, and Cyst-Like Structures

Abstract

The cyst stage of Entamoeba histolytica is a promising therapeutic target against human amoebiasis. Our research team previously reported the production in vitro of Cyst-Like Structures (CLS) sharing structural features with cysts, including rounded shape, size reduction, multinucleation, and the formation of a chitin wall coupled to the overexpression of glucosamine 6-phosphate isomerase, the rate-limiting enzyme of the chitin synthesis pathway. A proteomic study of E. histolytica trophozoites, cysts, and in vitro-produced CLS is reported herein to determine the nature of CLS, widen our knowledge on the cyst stage, and identify possible proteins and pathways involved in the encystment process. Total protein extracts were obtained from E. histolytica trophozoites, CLS, and partially purified cysts recovered from the feces of amoebic human patients; extracts were trypsin-digested and analyzed by LC-MS/MS. In total, 1029 proteins were identified in trophozoites, 550 in CLS, and 411 in cysts, with 539, 299, and 84 proteins unique to each sample, respectively, and only 74 proteins shared by all three stages. About 70% of CLS proteins were shared with trophozoites, even though differences were observed in the relative protein abundance. While trophozoites showed a greater abundance of proteins associated to a metabolically active cell, CLS showed higher expression of proteins related to proteolysis, redox homeostasis, and stress response. In addition, the expression of genes encoding for the cyst wall proteins Jessie and Jacob was detected by RT-PCR and the Jacob protein identified by Western blotting and immunofluorescence in CLS. However, the proteomic profile of cysts as determined by LC-MS/MS was very dissimilar to that of trophozoites and CLS, with almost 40% of hypothetical proteins. Our global results suggest that CLS are more alike to trophozoites than to cysts, and they could be generated as a rapid survival response of trophozoites to a stressful condition, which allows the parasite to survive temporarily inside a chitin-like resistant cover containing Jacob protein. Our findings lead us to suggest that encystment and CLS formation could be distinct stress responses. In addition, we show that cysts express a high number of genes with unknown function, including four new, highly antigenic, possibly membrane-located proteins that could be targets of therapeutic and diagnostic usefulness.

Membrane Trafficking Modulation during Entamoeba Encystation

“…However, two proteomic analyses of E . histolytica have identified several membrane trafficking proteins (among others) present in the cyst 63 , 64 , suggesting that at least some MTS proteins are retained.…”

“…Proteomics analyses of E . histolytica cysts have shown a handful of MTS proteins, among many others, present in mature cysts 63 , 64 , raising the possibility that these mRNAs may indeed be translated. Two proteins identified were clathrin heavy chain and a TBC-B homologue (EHI_094140).…”

Proteomic Insights into the Biology of the Most Important Foodborne Parasites in Europe

“…Amoebiasis is a disease caused by the protozoan parasite Entamoeba histolytica ( E. histolytica ). When a person swallows something contaminated with E. histolytica cysts (water or food products), infection may occur [ 61 ]. Only about 10% to 20% of people who are infected with E. histolytica will become ill due to the infection.…”

Prevalence of amoebic dysentery and giardiasis in Al-Najaf

“…This parasite is extant in polluted water and sludge in 2 forms: trophozoites and cysts (Begum et al, 2015) . The life cycle of it initiate with cysts entry to the body by drinking water or eating food which impure or contaminated with feces while the trophozoites appear after excystation in small intestine then belay itself the large intestine by clinging to the colonic mucosa (Begum et al, 2015;Luna-Nácar et al, 2016) leading to Different severe symptoms of which the ulcer, which cause intestinal colic and diarrhea and bleeding of mucosa, loss of appetite, vomiting (Kucik et al, 2004) , the main source of infection that leading to anemia and bad nutrition anemia is the lack of sodium and potassium ions (Keen, 2013) .…”