Emetine-resistant mutants of Entamoeba histolytica overexpress mRNAs for multidrug resistance.
We report here the identification of multiple 344-bp segments of amoeba DNA similar to the human multidrug resistance (mdr) gene using primers to conserved regions of the P-glycoprotein and the polymerase chain reaction (PCR). The amino acid sequences of amoeba mdr-like PCR products were 46-97% identical to each other, 46-51% identical to human mdr1 sequences, and 30-35% identical with Plasmodium falciparum mdr-like sequences. On Northern blots, the mdr-like PCR products identified amoeba mRNAs 4.5-5 kb long, similar to the 5-kb mRNAs reported for the mammalian mdr gene. These mdr-like mRNAs were increased at least 7 times in emetine-resistant mutant clone C2 amoebae vs. wild-type clone A parasites. Furthermore, the expression of the mdr-like mRNAs was increased 3-4 times when clone C2 mutants were grown under drug pressure vs. the same parasites grown without emetine. In contrast, the number of genomic copies of the mdr-like DNA segments was not increased in the mutant clone C2 vs. the wild-type clone A amoebae, and no rearrangements of the mdr-like DNA segments by the mutant were identified on Southern blots. In conclusion, there appears to be a family of mdr-like genes in Entamoeba histolytica, which may be involved in drug resistance by the parasite because they are overexpressed in drug-resistant mutants.
Samuelson
J
J
Department of Tropical Public Health, Harvard School of Public Health, Boston, MA 02115.
Ayala
P
P
Orozco
E
E
Wirth
D
D
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004351:Drug Resistance / Q000235:genetics*; D004640:Emetine / Q000494:pharmacology*; D004748:Entamoeba histolytica / Q000187:drug effects* / Q000235:genetics; D005784:Gene Amplification; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D008562:Membrane Glycoproteins; D004640:Emetine