DEVELOPMENTAL GENE-EXPRESSION IN LEISHMANIA-DONOVANI - DIFFERENTIAL CLONING AND ANALYSIS OF AN AMASTIGOTE-STAGE-SPECIFIC GENE

Leishmania protozoans are the causative agents of leishmaniasis, a maj or parasitic,disease in humans. During their life cycle, Leishmania pr otozoans exist as flagellated promastigotes in the sand fly vector and as nonmotile amastigotes in the mammalian hosts. The promastigote-to- amastigote transformation occurs in the phagolysosomal compartment of the macrophage cell and is a critical step for the establishment of th e infection. To study this cytodifferentiation process, we differentia lly screened an amastigote cDNA library with life cycle stage-specific cDNA probes and isolated seven cDNAs representing amastigote-specific transcripts. Five of these were closely related (A2 series) iind reco gnized, by Northern (RNA) blot analyses, a 3.5-kb transcript in amasti gotes and in amastigote-infected macrophages. Expression of the amasti gote-specific A2 gene was induced in promastigotes when they were tran sferred from culture medium at 26 degrees C and pH 7.4 to medium at 37 degrees C and pH 4.5, conditions which mimic the macrophage phagolyso somal environment. A2 genes are clustered in tandem arrays, and a 6-kb fragment corresponding to a unit of the cluster,vas cloned and partia lly sequenced. An open reading frame found within the A2-transcribed r egion potentially encoded a 22-kDa protein containing repetitive seque nces. The recombinant A2 protein produced in Escherichia coil cells wa s specifically recognized by immune serum from a patient with visceral leishmaniasis. The A2 protein repetitive element has strong homology with an S antigen of Plasmodium falciparum, the protozoan parasite res ponsible for malaria. Both the A2 protein of Leishmania donovani and t he S antigen of P. falciparum are stage specific and developmentally e xpressed in mammalian hosts.