Mutant/mutation
In the 'promoter-swap' mutant the promoter of rab11a replaced by the promoter of the asexual blood stage gene clag (PBANKA_1400600, RhopH1A, cytoadherence linked asexual protein). This promoter is silent in gametocytes/ookinetes. In addition the mutant expresses GFP under the constitutive eef1a promoter.

Protein (function)
Endocytosis-mediated protein transport is generally necessary for the establishment and maintenance of polarity in epithelial cells and neurons, and the small GTPase RAB11A is an important regulator of protein transport via recycling endosomes. PbRAB11A is essential in blood stage asexual of Plasmodium.
Rab genes encode a subgroup of small GTP-binding proteins within the ras super-family that regulate targeting and fusion of transport vesicles within the secretory and endocytic pathways. In the genome of P. falciparum 11 genes have been identified that encode Rab GTPases. Rab11A is highly conserved in apicomplexan parasites. Unsuccessful attempts to disrupt Rab11A (RMgm-298) indicates an essential rol of Rab11A during asexual blood stage development.

Phenotype
A severely impaired zygote to ookinete development by >98%. Zygotes of both promoter swap lines remained spherical and failed to develop the characteristic elongated morphology of a wild type ookinete. Strongly reduced oocyst production. Wild type showed normal oocyst development in mosquito midguts [median 150, mean 227 oocysts per midgut (n = 3)] while pclag::rab11a (maximum of 4 oocysts per midgut with small size, mean 0.269, median 0 and n = 3). No sporozoite formation

Additional information
A polyclonal antibody raised against the C-terminal end of PBRab11A was used in Western analysis and demonstrated expression of pbrab11a across mixed asexual stages, schizonts, gametocytes and ookinetes. A smaller, ~23kDa band of unknown significance was detected indicating possible differential regulation or N-terminal processing of PbRAB11A in schizonts and mature ookinetes. Immunofluorescence microscopy using the same polyclonal antibody on fixed  parasites showed expression of PbRAB11A in mature schizonts (individual merozoites), 6h zygotes, ookinetes, oocysts as well as midgut sporozoites. In merozoites, oocysts (day 10) and midgut sporozoites (day 10), the localization of PbRAB11A appeared to be cytoplasmic. In the 6h zygote, PbRAB11A was cytoplasmic and appeared punctate while in the ookinete PbRAB11A was more peripheral and with a focus at the apical tip suggesting a role of PbRAB11A at the apical complex

Evidence is presented that:
- PbRAB11A is not necessary for the generation and deposition of the apical complex and the IMC.

From the abstract:
'The approach revealed that lack of PbRAB11A had no effect on gamete production and fertility rates however, the zygote to ookinete transition was almost totally inhibited and transmission through the mosquito was prevented. Lack of PbRAB11A did not prevent meiosis and mitosis, nor the establishment of polarity as indicated by the correct formation and positioning of the Inner Membrane Complex (IMC) and apical complex. However, morphological maturation was prevented and parasites remained spherical and immotile and furthermore, they were impaired in the secretion and distribution of microneme cargo. The data are consistent with the previously proposed model of RAB11A endosome mediated delivery of plasma membrane in Toxoplasma gondii if not its role in IMC formation and implicate it in microneme function'.

Other mutants