Eukaryotic Cell
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Eukaryotic Cell, October 2007, p. 1758-1765, Vol. 6, No. 10
1535-9778/07/$08.00+0     doi:10.1128/EC.00241-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Transcriptional Gene Silencing Reveals Two Distinct Groups of Entamoeba histolytica Gal/GalNAc-Lectin Light Subunits{triangledown}

Rivka Bracha, Yael Nuchamowitz, Nomy Wender, and David Mirelman*

Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel

Received 4 July 2007/ Accepted 20 August 2007

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.

* Corresponding author. Mailing address: Weizmann Institute of Science, Department of Biological Chemistry, PO Box 26, Rehovot 76100, Israel. Phone: 972-8-9344511. Fax: 972-8-9344118. E-mail: david.mirelman{at}weizmann.ac.il

{triangledown} Published ahead of print on 31 August 2007.

Eukaryotic Cell, October 2007, p. 1758-1765, Vol. 6, No. 10
1535-9778/07/$08.00+0     doi:10.1128/EC.00241-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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