Mutant/mutation
A double gene replacement mutant: In the mutant the P. berghei cpα and cpβ gene are replaced with the complete P. falciparum cpα and cpβ orthologs. The P. falciparum genes are under the control of the P. berghei cpα and cpβ promoter region, respectively. The mutant has been generated by crossing the single gene cpα replacement mutant RMgm-1154 with  the single gene cpβ replacement mutant RMgm-1156 in mosquitoes and selection of the double replacment mutant

Protein (function)
One of the few conserved actin-binding proteins of Plasmodium parasites is the F-actin capping protein (CP), which is found in all eukaryotic organisms and metazoan cell types

CP binds in a calcium-independent manner to the fast growing (barbed) ends of F-actin, thereby blocking subunit exchange. CP also belongs to the defined set of proteins that are needed to reconstitute actin-based motility in vitro.

Active CP is composed of two subunits, CPα and CPβ, and production of recombinant active CP in Escherichia coli (E. coli) is typically only achieved by co-expression of both subunits

Plasmodium CPβ is encoded by a single open reading frame, whereas CPα is composed of nine small exons. Overall, Plasmodium CPα-subunits share approximately 19% amino acid sequence identity with other eukaryotic CPα-subunits, and 50-90% identity across different Plasmodium species.

The residues that contribute to actin binding and heterodimer formation are conserved.

CPβ-subunit of rodent malaria parasite P. berghei (PbCPβ) as an essential regulator of sporozoite motility and malaria transmission. Deletion of PbCPβ did not influence asexual and sexual blood-stage development in the mammalian host. In the insect vector, Anopheles mosquitoes, mutant parasites displayed defective motility, which completely arrested life cycle progression at the sporozoite stage. It has been shown that recombinant P. berghei CPα/β heterodimers display capping activity on heterologous non-muscle actin. The stage-specific function of CPβ in sporozoites implies that CPα alone might be functional during blood infection of cpβ(-) parasites. Given that independent functions of CP subunits have not been described, this notion was unexpected and prompted to investigate the cellular role(s) of Plasmodium CPα for parasite life cycle progression.

In this study it is shown that the two CP subunits can be functionally separated. Unlike the beta subunit, the CPalpha subunit of the apicomplexan parasite Plasmodium is refractory to targeted gene deletion during blood infection in the mammalian host.

See the link F-actin-capping protein subunits for other mutants with mutated F-actin-capping protein subunits

Phenotype
Phenotype analyses of the single gene cpα replacement mutant RMgm-1154 indicate that P. falciparum CPα can fully complement P. berghei CPα during blood stage development. However, the mutant shows strongly reduced numbers of oocysts in A. stephensi mosquitoes and absence of salivary gland sporozites.

Phenotype analyses of with  the single gene cpβ replacement mutant RMgm-1156  show strongly reduced numbers of oocysts in A. stephensi mosquitoes and absence of salivary gland sporozites.

The double replacement P. berghei mutant in which both the cpα and cpβ genes are replaced with the complete P. falciparum cpα and cpβ genes (RMgm-1157) shows a partial rescue of the phenotype with low numbers of sporozoites present in salivary glands.

Combined these observations indicate that i) trans-species complementation of both subunits is needed to rescue the severe sporozoite defect as seen in both single replacement mutants and that ii) a functional CP heterodimer is necessary for normal oocyst and sporozoite development.

Additional information

Other mutants
See the link F-actin-capping protein subunits for other mutants with mutated F-actin-capping protein subunits