Transcriptional gene silencing reveals two distinct groups of Entamoeba histolytica Gal/GalNAc-lectin light subunits.
The Entamoeba histolytica cell surface Gal/GalNAc-inhibitable lectin is a heterodimer between a heavy (170 kDa) subunit linked via a disulfide bond to a light (31 to 35 kDa) subunit. Five light subunit genes with high homology have been identified (Ehlgl1 to -5). We have previously shown that silencing of the expression of Ehlgl1, in the G3 trophozoites which had already been silenced in the amoebapore gene (Ehap-a), also suppressed the transcription of Ehlgl2 and -3 (strain RBV). The total absence of the lgl1 to -3 subunits in the RBV trophozoites affected their ability to cap the surface Gal-lectin molecules to the uroid region. We have now found that in the RBV trophozoites, the lgl4 and -5 subunits (31 kDa) are overexpressed and appear to compensate for the missing lgl1 to -3 in the heterodimer complex. Transcriptional silencing of Ehlgl5 was achieved by transfection of G3 trophozoites with a plasmid containing the open reading frame of Ehlgl5 ligated to the 5' promoter region of the Ehap-a gene. The transfected trophozoites (strain L5) were silenced in Ehlgl5 and the closely related Ehlgl4, while the expression of the larger lgl1 to -3 subunits was upregulated. L5 trophozoites retained their ability to cap the Gal-lectin molecules. Attempts to simultaneously silence all of the Ehlgl genes have failed so far, possibly due to their crucial importance to the Gal-lectin functions. Our ability to silence part of the genes belonging to the same family can serve as a tool to study the relationships and functions of the members of other gene families.
Bracha
Rivka
R
Weizmann Institute of Science, Department of Biological Chemistry, Rehovot, Israel.
Nuchamowitz
Yael
Y
Wender
Nomy
N
Mirelman
David
D
D000818:Animals; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D004748:Entamoeba histolytica / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D017125:Genes, Protozoan / Q000235:genetics; D006801:Humans; D037102:Lectins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010271:Parasites / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D010587:Phagocytosis; D010641:Phenotype; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D014158:Transcription, Genetic; D053058:Trophozoites / Q000378:metabolism
D037102:Lectins; D021122:Protein Subunits; D015800:Protozoan Proteins