Eukaryotic Cell
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Bracha, R.
Right arrow Articles by Mirelman, D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Bracha, R.
Right arrow Articles by Mirelman, D.
Eukaryotic Cell, April 2003, p. 295-305, Vol. 2, No. 2
1535-9778/03/$08.00+0     DOI: 10.1128/EC.2.2.295-305.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Transcriptional Silencing of an Amoebapore Gene in Entamoeba histolytica: Molecular Analysis and Effect on Pathogenicity

Rivka Bracha, Yael Nuchamowitz, and David Mirelman*

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

Received 17 October 2002/ Accepted 17 January 2003

Transcriptional silencing of the gene coding for amoebapore A (AP-A) was observed when trophozoites of Entamoeba histolytica were transfected with a hybrid plasmid construct containing the ap-a gene flanked by the upstream and downstream segments of the original Ehap-a gene. Transfectants were totally devoid of ap-a transcript and AP-A protein. An identical silencing effect was observed upon transfection with a plasmid that contained only the 5' upstream region of ap-a. Removal of the selecting antibiotic enabled the isolation of plasmidless clones, which retained in their progeny the silenced phenotype. E. histolytica cells were able to overexpress ap-a when transfected with a plasmid containing the gene flanked by the 5' and 3' regions of the EhRP-L21 gene. This plasmid, however, could not express ap-a in the retransfected, cloned trophozoites lacking AP-A. This is the first report of gene silencing in E. histolytica, and the mechanism appears to belong to transcriptional gene silencing and not to posttranscriptional gene silencing. This conclusion is based on the following results: (i) silencing was achieved by transfection of homologous 5' flanking sequences (470 bp of the Ehap-a gene), (ii) transcription initiation of Ehap-a was found to be blocked, and (iii) short double-stranded RNA fragments of the ap-a coding and noncoding sequences were not detected. Trophozoites lacking AP-A are nonpathogenic and impaired in their bacteriolytic capability.

* Corresponding author. Mailing address: Department of Biological Chemistry, Weizmann Institute of Science, P.O. Box 26, Rehovot 76100, Israel. Phone: 972-8-9344511. Fax: 972-8-9468256. E-mail: david.mirelman{at}weizmann.ac.il.

Eukaryotic Cell, April 2003, p. 295-305, Vol. 2, No. 2
1535-9778/03/$08.00+0     DOI: 10.1128/EC.2.2.295-305.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:



HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Appl. Environ. Microbiol. Infect. Immun. J. Bacteriol.
Mol. Cell Biol. Microbiol. Mol. Biol. Rev. ALL ASM JOURNALS
Copyright © 2003 by the American Society for Microbiology.