Proteomic Study of Entamoeba histolytica Trophozoites, Cysts, and Cyst-Like Structures
Por:
Luna-Nacar, Milka, Navarrete-Perea, Jose, Moguel, Barbara, Bobes, Raul J., Laclette, Juan P., Carrero, Julio C.
Publicada:
26 may 2016
Resumen:
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.
Filiaciones:
Luna-Nacar, Milka:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
Navarrete-Perea, Jose:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
Moguel, Barbara:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
Bobes, Raul J.:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
Laclette, Juan P.:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
Carrero, Julio C.:
Univ Nacl Autonoma Mexico, Inst Invest Biomed, Dept Immunol, Tercer Circuito Exterior S-N,Ciudad Univ, Mexico City 04510, DF, Mexico
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