Molecular cloning and characterization of a novel acidic microneme protein ( Etmic-2) from the apicomplexan protozoan parasite, Eimeria tenella

Abstract

A 50 kDa acidic protein, which is found within the microneme organelles of Eimeria tenella sporozoites and merozoites and called E. tenella mic-2, was cloned by immunoscreening of a cDNA expression library. The expression of the protein and its mRNA during the developmental cycle of the parasite was consistent with de novo formation of microneme organelles during both sporulation and schizogony. Although micronemal in origin, indirect immunofluorescent antibody labelling on gluraraldehyde fixed parasites, indicated that the protein was translocated to the sporozoite surface, and, during host cell invasion the protein was focussed at the point of parasite entry and secreted from the host-parasite interface. Either during or just after invasion, Etmic-2 protein became transiently dispersed over the entire surface of the infected cell. One hour after adding sporozoites to host cells, no detectable Etmic-2 protein remained on the host cell surface. A full length cDNA corresponding to Etmic-2 predicted a protein with a classical signal peptide that preceded the mature N-terminus of the protein as determined by direct microsequencing. Regions of the Etmic-2 protein have highly significant similarities to regions within Drosophila melanogaster tropomyosin II and within two known substrates of the cellular regulatory enzyme protein kinase C.