Mutant/mutation
In the promoter swap' mutant: the promoter of the endogenous gap45  gene is replaced by the promoter of ama1 (PBANKA_091500).

Protein (function)
GAP45 is part of the actomyosin motor located under the surface membrane of the invasive/motile stages of the parasite. In this motor, actin filaments attached to membrane proteins bind an atypical myosin (MyoA), which is in turn attached by its C-terminal end to a sub-pellicular structure called the inner membrane complex (IMC) via a myosin light chain (or myosin A tail domain interacting protein, MTIP). MyoA, MTIP and the 45 kDa glideosome associated protein (GAP45) cotranslationally form a complex, which interacts with other proteins such as GAP50 at the IMC.

Phenotype
GAP45 is considered vital for blood stage development/multiplication (merozoite invasion of erythrocytes).
In the 'promoter-swap' mutant the promoter of gap45  is replaced by an 'asexual blood stage’ promoter that is silent (or has a strongly reduced activity) in ookinetes (the promoter of ama1, PBANKA_091500). Expression of GAP45 was absent in mutant ookinetes whereas wild type ookinetes express GAP45.

Phenotype analyses of the promoter swap mutant indicate that GAP45 has an essential role in ookinete development. Mutant parasites showed normal gamete formation and fertilization. However, zygotes failed to develop into ookinetes and remained spherical.

Additional information
Electron microscope analyses of zygotes 20 hr after fertilization showed an intact apical complex featuring an apical collar, inner membrane complex, microtubules, and micronemes. However, these structures were detached from the plasmalemma and were ‘‘free floating’’ in the cytoplasm of a round zygote.

The mutant was complemented by additionally inserting a cassette expressing P. falciparum GAP45 from the P. berghei gap45 promoter into the Pama1-gap45 promoter swap construct. Genomic integration of this allele at the gap45 locus led to the expression of Pfgap45 mRNA and protein in schizonts and ookinetes. Complementation restored the formation of ookinetes, which were fully motile (data not shown), confirming that the unexpected block in zygote development is a true loss of function phenotype of gap45

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