Mutant/mutation
The mutant lacks expression of PLP1 (SPECT2). and expresses GFP under the control of the HSP70 promoter.  The parasites have been generated and selected using the GOMO method of transfection (see RMgm-1026 for more details for this transfection method and the construct used).

Protein (function)
The spect2/pplp1 gene is a member of a small, conserved family of proteins encoding perforin-like proteins containing membrane-attack complex/perforin domains (MACPF). The P. berghei genome contains 5 PLP proteins

Phenotype
Mutant sporozoites were motile and developed into EEFs in vitro (in HepG2/CD81 cells) as efficiently as control parasites, but were poorly infective to mice in vivo, especially when administered through mosquito bites, the natural transmission route. Evidence is presented that sporozoites form transient vacuoles (TVs) in traversed hepatocytes and that mutant sporozoites fail to egress from TVs, indicating that PLP1 is required for sporozoite egress from nonreplicative TVs, but not for entry into cells.

Additional information
Sporozoites migrate from the dermis to the liver, where they invade hepatocytes through a moving junction (MJ) to form a replicative parasitophorous vacuole (PV). Sporozoites need to traverse cells during progression through host tissues, a process requiring parasite perforin-like protein 1 (PLP1). In the paper evidence is presented that i) sporozoites traverse cells inside transient vacuoles that precede PV formation; ii) sporozoites initially invade cells inside transient vacuoles by an active MJ-independent process that does not require vacuole membrane remodeling or release of parasite secretory organelles typically involved in invasion; iii) sporozoites use pH sensing and PLP1 to exit these vacuoles and avoid degradation by host lysosomes; iv parasites enter the MJ-dependent PV, which has a different membrane composition, precluding lysosome fusion.

The mutant parasites are selected by a combination of positive selection (pyrimethamine), negative selection (5-FC) and FACS sorting (GOMO transfection; 'Gene Out Marker Out'; see mutant RMgm-1026 for more details for this transfection method and the construct used).

Other mutants
See the link PPLP for other mutants with disrupted or tagged pplp genes