Like all members of Apicomplexa, the motile form of Plasmodium, the merozoite, possesses an
elaborate apical complex composed of 3 organelles; micronemes, rhoptries and dense granules.
Invasion of erythrocytes by Plasmodium merozoites is a tightly regulated multi-step process that
begins with the cooperation of proteins released from micronemes and rhoptries to mediate
attachment of parasites to the host cell membrane and to establish the parasitophorous vacuole.
RhopH complex is a very abundant rhoptry secretion composed of 3 protein members; RhopH1,
RhopH2 and RhopH3. RhopH1 is represented by either Clag2, Clag3.1, or Clag9. RhopH2 was
found to traffic to the rhoptries via its first 24 amino acids. To further address how the other
members of RhopH complex proteins are trafficked to rhoptries, the present study established a
new transgenic Plasmodium falciparum line expressing a large chimeric protein containing
almost ¾ of Clag3.1 fused to the N-terminus of GFP via a linker region under the control of
rhoph2 promoter and its signal peptide encoding sequence. The data showed that this chimera
was expressed efficiently indicating the strength of rhoph2 promoter, however, the chimera was
trapped in the early secretory pathway and did not reach rhoptries. Provided a correct folding of
this protein, these data could imply that the middle part of Clag3.1 (aa 484-1064) contain some
information that interfere with the targeting contained in the first 24 amino acids of RhopH2.
The C-terminus of Clag3.1 could contain Clag3.1-specific rhoptry targeting information. Based
on literature available in the international databases, this is the first report to successfully
express such a large rhoptry chimeric GFP protein.