The Plasmodium falciparum RhopH2 promoter and first 24 amino acids are sufficient to target proteins to the rhoptries.
The rhoptry secretory organelles of the malaria parasite, Plasmodium falciparum, contain a RhopH complex, which is composed of the proteins RhopH1, RhopH2, and RhopH3. RhopH1 is encoded by the rhoph1/clag multi-gene family, whereas RhopH2 and RhopH3 are encoded by single-copy genes. The precise function of the RhopH complex has not been identified, but it has been shown that the component proteins are involved in erythrocyte binding and perhaps participate in the formation of the parasitophorous vacuolar membrane. In this study, we have isolated pfrhoph2 promoter plus the signal peptide encoding sequence and generated transgene expression constructs to evaluate a trafficking and the RhopH complex formation in transgenic P. falciparum parasite lines. Interestingly, we found that the N-terminal 24 amino acids of RhopH2, including signal peptide sequence, were sufficient to target GFP to the rhoptries under the rhoph2 promoter. Because it was previously shown that the timing of the expression alone could not target proteins to the apical organelles, this targeting is likely mediated via a unique mechanism that is dependent on N-terminal 24 amino acids of RhopH2 early in the secretory pathway. The N-terminal one third of Clag3.1, which contains a distinct conserved domain with Toxoplasma gondii RON2, can not associate the RhopH complex as a GFP chimera, but a c-Myc-Clag3.1 chimera lacking the C-terminus successfully associates the RhopH complex indicating that cooperation of middle region is likely required but the C-terminus is not necessary.
Ghoneim
Ahmed
A
Department of Molecular Parasitology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
Kaneko
Osamu
O
Tsuboi
Takafumi
T
Torii
Motomi
M
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D015153:Blotting, Western; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000737:chemistry* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D021382:Protein Sorting Signals; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C502072:cytoadherence-linked asexual protein 3, Plasmodium falciparum; C073168:rhoptry-associated antigen-2, Plasmodium; D049452:Green Fluorescent Proteins