Mutant/mutation
The mutant expresses a C-terminal GFP-tagged version of LIMP.

Protein (function)
The gene comprises six exons and five introns (this organisation is conserved throughout the genus) and encodes a protein of 110 amino acids (aa) with a 22 aa long signal peptide and a molecular weight of 13 kDa. Ab initio protein structure predictions  indicate that LIMP (I23 to G110) consists of three beta sheets opposed to two a-helices. LIMP is highly conserved among the various Plasmodium species; similarly short proteins are present in related api-complexan parasites, where the homology is focused on a 22 amino acid proline-rich region adjacent to the signal peptide.

Phenotype
Analyses of mutants lacking expression of LIPM (RMgm-4165) indicate a role for LIMP during mosquito midgut colonisation by the ookinete, and a crucial function for the protein during salivary gland invasion, but not for development; neither ookinete nor sporozoite formation were affected by the absence of LIMP. Δlimp sporozoites are impaired in hepatocyte transmigration, adhesion and invasion and do not glide.

The analyses a GFP-tagged version of LIMP (RMgm-4166) showed the following:
'No protein expression in asexual stage parasites or gametocytes; limp::gfp is translated in the ookinete stage and its expression is visible in crystalloids (transient and putative storage organelles of the ookinete) and the surface; the protein produced a faint ‘dusting’ on the surface of midgut and salivary gland sporozoites. In agreement with previously published results limp mRNA was not detected in asexual blood stages; mRNA is however present in gametocytes (while protein is not) and in ookinetes (where protein is present); this is consistent with limp being translationally repressed and maternally provided to the developing ookinete. Western blot analysis of ookinete material confirmed expression of a GFP fusion protein of the expected size. Parasites expressing LIMP::GFP showed no defects in liver stage development in vitro nor during transmission to the rodent host by mosquito bite'.

Evidence is presented for:
- limp::gfp sporozoites  move significantly slower  despite limp::gfp salivary gland sporozoites producing normal numbers of CSP trails
- No evidence for shedding of LIMP during sporozoite migration
- LIMP::GFP parasites glide with a ‘limp’

GFP tagging of LIMP caused a limping defect during movement with reduced speed and transient curvature changes of the parasite.

The aberrant gliding motility was not observed in sporozoites expressing N-terminal mCherry-tagged LIMP (RMgm-4167) or C-terminal myc-tagged LIMP (RMgm-4168)

Additional information
Using an immuno-electron microscopy (immuno-EM) approach, we corroborated LIMP::GFP surface localisation in sporozoites in three independent staining experiments with three different parasite samples (using anti-GFP antibodies). Seventy-six percent of gold particles (from a total of 281) were associated with the parasite PM, 11% outside of parasites; 13% were intracellular although not clearly associated with any specific organelle.

Other mutants
See PF3D7_1207300