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J Biol Chem, Vol. 273, Issue 24, 15119-15124, June 12, 1998

Precise Timing of Expression of a Plasmodium falciparum-derived Transgene in Plasmodium berghei Is a Critical Determinant of Subsequent Subcellular Localization

Clemens H. M. Kocken, Anne Marie van der Wel, Martin A. Dubbeld, David L. Narum, Franciscus M. van de RijkeDagger , Geert-Jan van Gemert§, Xander van der Linde, Lawrie H. Bannister, Chris Janse**, Andrew P. Waters**, and Alan W. Thomas

From the Department of Parasitology, Biomedical Primate Research Centre, Lange Kleiweg 157, 2280 GJ Rijswijk, The Netherlands, the Dagger  Leiden University Medical Center, Department of Molecular Cell Biology, Laboratory of Cytochemistry and Cytometry, Wassenaarseweg 72, 2333 AL Leiden, The Netherlands, the § Department of Medical Microbiology, University of Nijmegen, Geert Groote Plein 24, 6500 HB Nijmegen, The Netherlands, the  Department of Anatomy, The Medical School, Guy's Hospital, London SE1 9RT, Great Britain, and the ** Department of Parasitology, Leiden University, Wassenaarseweg 62, 2300 RC Leiden, The Netherlands

The development of transfection technology for malaria parasites holds significant promise for a more detailed characterization of molecules targeted by vaccines or drugs. One asexual blood stage vaccine candidate, apical membrane antigen-1 (AMA-1) of merozoite rhoptries has been shown to be the target of inhibitory, protective antibodies in both in vitro and in vivo studies. We have investigated heterologous (trans-species) expression of the human malaria Plasmodium falciparum AMA-1 (PF83/AMA-1) in the rodent parasite Plasmodium berghei. Transfected P. berghei expressed correctly folded and processed PF83/AMA-1 under control of both pb66/ama-1 and dhfr-ts promoters. Timing of expression was highly promoter-dependent and was critical for subsequent subcellular localization. Under control of pb66/ama-1, PF83/AMA-1 expression and localization in P. berghei was limited to the rhoptries of mature schizonts, similar to that observed for PF83/AMA-1 in P. falciparum. In contrast the dhfr-ts promoter permitted PF83/AMA-1 expression throughout schizogony as well as in gametocytes and gametes. Localization was aberrant and included direct expression at the merozoite and gamete surface. Processing from the full-length 83-kDa protein to a 66-kDa protein was observed not only in schizonts but also in gametocytes, indicating that processing could be mediated outside of rhoptries by a common protease. Trans-species expressed PF83/AMA-1 was highly immunogenic in mice, resulting in a response against a functionally critical domain of the molecule.

Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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Copyright © 1998 by the American Society for Biochemistry and Molecular Biology.