Recent studies on the development of Plasmodium falciparum in the mature human erythrocyte, a quiescent cell lacking all organelles and machinery to synthesize proteins, has suggested the presence of an extensive protein secretory pathway. Plasmodium proteins are transported through ER/Golgi-like infrastructure within the parasite and, certain proteins are further exported from the parasitophorous vacuole membrane to the cytoplasm of erythrocyte through a tubovesicular network. To help define the protein secretory pathway in the malaria parasite at the molecular level, we have decided to investigate the role of the Rab proteins, a family of Ras-related GTP-binding proteins which control vesicular transport in all eukaryotic cell types examined thus far. We have identified several cDNA clones encoding Rab homologues from a Plasmodium falciparum lgt11 library. A PfRab1 protein, demonstrating extensive homology to the mammalian Rab1 and yeast YPT1, displays typical features of the Ras-like GTP-binding domains and a geranylgeranlyation motif on its C-terminus. Rab1 proteins in eukaryotes are ubiquitously expressed and directly involved in vesicular transport from the ER to cis Golgi compartment. In Northern blot analysis, a transcript of approx. 1.3 kb was detected in the sexual stage of gametes/zygotes, while little or no transcript was detected in the asexual erythrocytic stage of merozoites. To clarify and confirm this finding, immunofluorescence microscopy using rabbit anti-PfRab1 peptide antibody was performed. PfRab1 was detected predominantly in gametocytes (Stage III, IV and V), newly exflagellated gametes and 4-hr post-exflagellated gametes/zygotes. Weak fluorescence was detected in the asexual stages of late trophozoites, schizonts and the stage II of gametocytes. The fluorescence was detected only in the cytoplasm and not localized to any specific region. These results suggest that: 1) the asexual stages of parasite probably lack a specific Golgi compartment. Or, 2) If there is a Golgi-like compartment, either it rarely utilizes PfRab1 in the vesicular transport between ER and Golgi or another member of the Rab family functions in place of PfRab1. We have also expressed Pfhsp60 in E. coli and used to raise specific antibody in rabbits. Pfhsp60 will be further investigated for its potential as an adjuvant or immunogen in the form of a naked DNA vaccine.
