A homologue of N-ethylmaleimide-sensitive factor in the malaria parasite Plasmodium falciparum is exported and localized in vesicular structures in the cytoplasm of infected erythrocytes in the brefeldin A-sensitive pathway

N-Ethylmaleimide-sensitive factor (NSF) and its homologues play a central r ole in vesicular trafficking in eukaryotic cells. We have identified a NSF homologue in Plasmodium falciparum (PfNSF), The reported PfNSF gene sequenc e (GenBank(TM) accession number CAB10575) indicated that PfNSF comprises 78 3 amino acids with a calculated molecular weight of 89,133, The overall ide ntities of its gene and amino acid sequences with those of rat NSF are 50.9 and 48.8%, respectively. Reverse transcription-polymerase chain reaction a nalysis and Northern blotting with total P, falciparum RNA indicated expres sion of the PfNSF gene. Polyclonal. antibodies against a conserved region o f NSF specifically recognized an 89-kDa polypeptide in the parasite cells. After homogenization of the parasite cells, similar to 90% of an 89-kDa pol ypeptide is associated with particulate fraction, suggesting membrane-bound nature of PfNSF. PfNSF was present within both the parasite cells and the vesicular structure outside of the parasite cells, The export of PfNSF outs ide of the parasite cells appears to occur at the early trophozoite stage a nd to terminate at the merozoite stage. The export of PfNSF is inhibited by brefeldin A, with 9 muM causing 50% inhibition. Immuno-electromicroscopy i ndicated that intracellular PfNSF was associated with organelles such as fo od vacuoles and that extracellular PfNSF was associated with vesicular stru ctures in the erythrocyte cytoplasm, These results indicate that PfNSF expr essed in the malaria parasite is exported to the extracellular space and th en localized in intraerythrocytic vesicles in a brefeldin A-sensitive manne r. It is suggested that a vesicular transport mechanism is involved in prot ein export targeted to erythrocyte membranes during intraerythrocytic devel opment of the malaria parasite.