Mutant/mutation
The mutant lacks expression of PPKL (protein phosphatase containing kelch-like domains) and expresses GFP under control of the constitutive eef1a promoter.

Protein (function)
PPKL is a distinctive PP1-related enzyme belonging to the PPP family of phosphatases comprising an N-terminal kelch repeat domain and a C-terminal PP1-like phosphatase domain (named PPKL: Protein Phosphatase with Kelch-Like domains). It has been detected in the apicomplexans Cryptosporidium hominis, Toxoplasma gondii and Theileria parva (one gene per genome), as well as in the land plants Arabidopsis thaliana and Oryza sativa (4 and 5 genes respectively). The kelch motif is widespread and involved in many cellular functions, particularly in actin-based cytoskeleton formation and transcriptional regulation. PPKL has only been studied in detail in Arabidopsis thaliana (where it is known as BSU1 or bri1 suppressor1) and along with the kinase BIN2, is involved in brassinosteroid hormone signalling and phosphorylation of the transcription factors BZR1 and BES1

In contrast to the human phosphatome (comprising approximately 156 phosphatases the Plasmodium genome codes for one of the smallest phosphatomes of all the eukaryotic phyla known to date, with 27 putative protein phosphatases falling into four major classes: phosphoprotein phosphatases (PPPs), metallo-dependent protein phosphatases (PPMs), protein tyrosine phosphatases (PTPs) and NLI interacting factor-like phosphatases (NIFs).

Phenotype
Phenotype analyses indicate an essential role of PPKL in maturation of ookinetes.
Deletion of the ppkl gene caused abnormal ookinete development, loss of ookinete motility and failure of ookinetes to invade the mosquito midgut epithelium. Evidence is presented for aberrant formation of the apical end and disorganization of microtubules.

Additional information
Pbppkl was disrupted in 2 independent experiments and transgenic parasites were generated in both non-fluorescent background (P. berghei ANKA) and a wild type line which constitutively expresses a gfp transgene under the control of the eef1a promoter (P. berghei ANKA 507cl1) described here.

While, ppkl mRNA was found in all asexual blood stages, gametocytes and ookinetes, PPKL protein expression was detected only in schizonts, female gametocytes and ookinetes (see also mutants RMgm-786 and RMgm-788 that express an endogenous GFP-tagged version of PPKL). Immunofluorescence studies using anti-PPKL polyclonal antibody showed expression in merozoites, female gametocytes, zygotes, ookinete, and sporozoites. Only female gametocytes and not males expressed PPKL. The phosphatase is localized predominantly in the cytoplasm with a particularly intense expression at the apical tip of the developing and mature ookinete.

Since the effect of PPKL absence was visible post-fertilization it was examined whether the defect was male or female specific by performing genetic crosses between ppkl– parasites and parasites lines which are specifically male fertile (line 270, p47-) or female fertile (line 137, p48/45-). Consistent with the observed wild type pattern of PPKL expression only crosses with wild type females from p48/45- (line 137) generated normal ookinetes (26%) revealing the requirement of PPKL to be female specific.

Other mutants
RMgm-786: A mutant expressing an endogenous GFP-tagged version of PPKL
RMgm-787: An independent mutant lacking expression of PPKL
RMgm-788: An independent mutant expressing an endogenous GFP-tagged version of PPKL