Mutant/mutation
The mutant expresses an additional copy of a (N-terminal) GFP-tagged copy of Actin I. The endogenous actin I gene is not tagged. The (N-terminal) GFP tagged actin I gene is introduced into the genome into the (silent) c/d-ssuRNA gene locus (by single cross-over integration). The GFP-tagged copy is under control of the constitutive eef1α promoter.

Protein (function)
Actin, a cytoskeletal protein, has many diverse functions in eukaryotic cells ranging from roles in cell motility, cell division, vesicle trafficking to functions in cell signaling and regulation of transcription. A critical property of actin is its ability to form filamentous polymers (F-actin), and a plethora of proteins are involved in the highly dynamic regulation of F-actin formation . Actins are highly conserved proteins that often exist in multiple isoforms in the eukaryotic cell and their expression is regulated both spatially and temporally during development. The number of conventional actin genes varies among eukaryotic organisms. A few single cell eukaryotes, such as Saccharomyces cerevisiae, Toxoplasma gondii, and Trypanosoma brucei encode a single actin gene, which results in lethality when targeted with gene ablation approaches. Many organisms, however, have several conventional actin genes.Apicomplexan parasites all encode one major actin isoform, here termed Actin I. All apicomplexan parasites also contain a number of actin-related and actin-like proteins. Plasmodium species species stand out in that they all encode a second conventional actin, termed Actin II.

Phenotype
In the mutant the GFP-tagged actin I is introduced into the genome as an additional copy and is under the control of the constitutive eef1α promoter. All different life cycle stages express GFP-tagged Actin I and was found to be broadly cytoplasmic. Sporozoites suggested a pellicular concentration of actin whereas ookinetes and merozoites showed more uniform fluorescence. No obvious dynamics of parasite actin could be detected in gliding or turning ookinetes.

Additional information
See also the paper of Kiamos et al. (2011. Mol. Biochem Parasitol.). In this paper filamentous actin was analysed in ookinetes using peptide antibodies against P. berghei Actin I. In this paper they report  tagging of Actin I tagged with YFP. The YFP-tagged acrtin I gene is under the control of the promoter of the circumsporozoite and TRAP-related protein (CTRP). No information is provided on the mode of tagging. They write '... detected the presence of actin in a well-defined region in the apical tip of the parasite ...a similar localization was seen in transgenic parasites, expressing as a second copy YFP-tagged actin I fusion protein......Fluorescently tagged actin may interfere with the normal function of endogenous actin; although not well understood, this effect may be dependent on the ratio of non-modified to modified form....We, therefore, hypothesize that the observed accumulation of YFP-actin I might reflect the fusion protein in the globular (G−) form. Clearly, we did not detect YFP-actin in a pattern consistent with F− actin.'

Other mutants