The developmental expression of Leishmania donovani A2 amastigote-specific genes is post-transcriptionally mediated and involves elements located in the 3'-untranslated region.
Leishmania donovani is a protozoan parasite that exists as a free-living promastigote in the sandfly insect vector and as an amastigote inside the mammalian host macrophage phagolysosome compartment. The L. donovani A2 genes have been described previously as developmentally expressed in amastigotes but can be induced experimentally in promastigotes by a combination of pH and temperature shifts, conditions that mimic the phagolysosomal compartment of the macrophage cell. Considering the importance of the amastigote stage in human infections, we have examined the molecular basis for amastigote stage-specific gene expression. Our results provide evidence that A2 developmental expression during the promastigote-to-amastigote cytodifferentiation is mediated through differential RNA stability and involves the A2 mRNA 3'-untranslated region. The site of processing in the 3'-untranslated region was a major factor for the accumulation of A2 mRNAs in cells incubated under phagolysosomal conditions. The stability of reporter gene transcripts bearing the A2 3'-untranslated region was increased in cells incubated at low pH, further confirming the importance of pH shift as an inducer for A2 expression. These observations contribute to defining the mechanism of amastigote-specific gene regulation in L. donovani. We also demonstrate the feasibility of using the A2 locus to express heterologous genes differentially in the amastigote form of the L. donovani parasite.
Charest
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Institute of Parasitology, McGill University, Ste. Anne de Bellevue, Québec, Canada H9X 3V9.
Zhang
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WW
Matlashewski
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G
D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers; D004175:Diptera / Q000469:parasitology; D018507:Gene Expression Regulation, Developmental; D017125:Genes, Protozoan; D006801:Humans; D007303:Insect Vectors; D007893:Leishmania donovani / Q000235:genetics / Q000502:physiology*; D008247:Lysosomes / Q000469:parasitology; D008264:Macrophages / Q000469:parasitology; D008322:Mammals; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010588:Phagosomes / Q000469:parasitology; D010957:Plasmids; D016133:Polymerase Chain Reaction; D012323:RNA Processing, Post-Transcriptional; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism*; D016053:RNA, Protozoan / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D014158:Transcription, Genetic
D017931:DNA Primers; D012333:RNA, Messenger; D016053:RNA, Protozoan; D011994:Recombinant Proteins