Plasmodium yoelii: Cloning and characterization of the gene encoding for the mitochondrial heat shock protein 60

Heat shock proteins are a highly conserved group of proteins required for t he correct folding, transport, and degradation of other proteins in vivo. T he Hsp70, Hsp90, and Hsp60 families are among the most widely studied famil ies. Hsp60 is found in eubacteria, mitochondria, and chloroplasts, where, i n cooperation with Hsp10, it participates in protein folding and translocat ion of proteins to the organelles. We have cloned and characterized the Hsp 60 gene of Plasmodium yoelii (PyHsp60). PyHsp60 is a single-copy gene, loca ted on chromosome 9, 10, or 11. The PyHsp60 cDNA sequence showed an open re ading frame of 1737 nucleotides that codes for a polypeptide of 579 amino a cids, with 93% amino acid identity to Plasmodium falciparum Hsp60 (PfHsp60) . Cloning and sequencing of a genomic PCP clone showed the presence of a 20 1-bp intron, located 141 bp downstream of the ATG codon. A single, heat-ind ucible, 2.3-kb transcript was detected in Northern blots of RNA isolated fr om blood stage parasites. Mouse antisera raised against a DNA vaccine vecto r that expresses PyHsp60 recognized sporozoites and liver- and blood-stage parasites by indirect fluorescent antibody test (IFAT). By Western blot, th ese antisera reacted with the mycobacterial Hsp65 and recognized a protein of approximately 65 kDa in P. yoelii sporozoites and P. falciparum blood st ages. These results show that PyHsp60 and PfHsp60 genes are homologous and that of the PyHsp60 gene encodes a heat-inducible, intracellular protein th at is expressed in several of the developmental stages of P. yoelii. (C) 19 99 Academic Press.