Mutant/mutation
The mutant expresses a C-terminal HA-tagged version of FLP and expresses GFP under control of a male and RFP under control of a female gametocyte specific promoter. This mutant does not contain a drug-selectable marker. The hdhfr/yfcu selectable marker has been removed from the genome by negative selection

Protein (function)
The P. berghei ferlin-like protein (FLP, PBANKA_1224400) contains six predicted C2 domains, a C-terminal transmembrane domain and a Fer domain (a ferlin-specific motif of unknown function). These typical features cluster FLP together with ferlin (PBANKA_1319300) into the ferlin protein family.Ferlins are typically embedded in vesicular membranes via a single, C-terminal transmembrane domain with the N-terminus of the protein facing the cytosol.' Negative attempts to disrupt FLP (RMgm-4576) indicates an essential role during growth/multiplication of asexual stages.
Ferlins are established mediators of calcium-induced membrane fusion and regulated exocytosis in higher eukaryotes

Phenotype
FLP is essential for blood stage development/multiplication (see RMgm-4576) for unsuccessful attempts to disrupt the flp gene). In the 'promoter-swap' mutant RMgm-4577 the promoter of flp is replaced by an 'asexual blood stage’ promoter that is silent (or has a strongly reduced activity) in gametocytes (the promoter of clag, PBANKA_1400600). Analyses of this mutant provide evidence that FLP plays a role in egress of male and female gametocytes from the host red blood cell after activation.

Analyses of the mutant FLP::HA, expressing HA-tagged FLP shows that:
The  FLP::HA  fusion  protein was detected in both schizont- and gametocyte-enriched blood stage lysates revealing the fusion protein at the expected size of 186 kDa. Antibody staining of fixed parasites confirmed expression in schizonts, showing a speckled signal. High abundance of  FLP::HA was observed in gametocytes, localizing to widespread, distinct intracellular speckles. Immuno-gold typically localized close to the vesicle membranes, which is in agreement  with  FLP being a predicted transmembrane protein.
Both male and female gametocytes showed a similar abundance and localization of FLP::HA. FLP-HA relocalizes to the cell peripery during activation of gametocytes.


Additional information
From the paper:
'To  assess  whether  FLP  co-localizes  with  these  previously  published  egress  factors G377 (PBANKA_1463000) and PPLP2 (PBANKA_1432400),  we  generated flp::HA;g377::mCherry  and  flp::HA;pplp2::mCherry  double  mutants,  in  which FLP is endogenously tagged with HA while G377 and PPLP2 are endogenously tagged with mCherry, respectively. Neither of the lines showed co-localization between FLP and either of the two proteins, arguing against FLP localization to the G377-positive OBs (osmiophilic bodies) or to the PPLP2-positive egress vesicles.' 
 
Other mutants