Transcriptional analysis of the EhPgp1 promoter of Entamoeba histolytica multidrug-resistant mutant.
We present here the cloning and characterization of the EhPgp1 multidrug resistance gene promoter isolated from the Entamoeba histolytica drug-resistant mutant clone C2. The EhPgp1 promoter lacks the typical TATA box and the transcriptional initiation sequences described for other E. histolytica promoters. The major transcription initiation site of the EhPgp1 gene was located at the ATG start codon. The EhPgp1 core promoter located within the first 244 base pairs showed a higher chloramphenicol acetyltransferase expression in the transfected trophozoites of clone C2 than in those of the sensitive clone A. Gel shift assays revealed three specific DNA-protein complexes (Ia, IIa, and IIIc) using nuclear extracts from clone C2, whereas three main complexes (If, IIf, and IIg) were limited to clone A. Competition assays suggested the presence of C/EBP-like and OCT-like proteins in complexes Ia and IIa, respectively, probably involved in the expression of the EhPgp1 gene, whereas complex IIIc was competed by GATA-1, C/EBP, OCT, and HOX oligonucleotides. Thus, differential DNA-protein complexes may be formed by transcriptional factors involved in the regulation of the EhPgp1 gene expression.
Gómez
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C
Department of Patología Experimental, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, CINVESTAV Instituto Politecnico Nacional, AP 14-740, Mexico 07300, Mexico.
Pérez
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DG
López-Bayghen
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Orozco
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D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D016054:DNA, Protozoan / Q000737:chemistry*; D018432:Drug Resistance, Multiple / Q000235:genetics*; D004748:Entamoeba histolytica / Q000187:drug effects / Q000235:genetics*; D005786:Gene Expression Regulation / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic
D016054:DNA, Protozoan; D014157:Transcription Factors