| Successful modification | The parasite was generated by the genetic modification |
| The mutant contains the following genetic modification(s) |
Gene tagging
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| Reference (PubMed-PMID number) |
Reference 1 (PMID number) : 31797966
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| MR4 number |
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| Parent parasite used to introduce the genetic modification |
| Rodent Malaria Parasite | P. berghei |
| Parent strain/line | P. berghei ANKA |
| Name parent line/clone |
Not applicable
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| Other information parent line | |
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| The mutant parasite was generated by |
| Name PI/Researcher | Angrisano F, Blagborough AM |
| Name Group/Department | Department of Life Sciences |
| Name Institute | Imperial College of Science, Technology and Medicine |
| City | London |
| Country | UK |
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| Name of the mutant parasite |
| RMgm number | RMgm-4711 |
| Principal name | PDI-Trans-GFP |
| Alternative name | |
| Standardized name | |
| Is the mutant parasite cloned after genetic modification | Yes |
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| Phenotype |
| Asexual blood stage | Immunofluorescence microscopy on non-permeabilized parasites confirmed PDI-Trans-GFP expression at the surface of activated male gametes and ookinetes. Live microscopy of mixed blood stages, female gametes and fixed immunofluorescence of sporozoites demonstrated that PDI-Trans-GFP is expressed across the entire parasitic lifecycle. |
| Gametocyte/Gamete | Immunofluorescence microscopy on non-permeabilized parasites confirmed PDI-Trans-GFP expression at the surface of activated male gametes and ookinetes. Live microscopy of mixed blood stages, female gametes and fixed immunofluorescence of sporozoites demonstrated that PDI-Trans-GFP is expressed across the entire parasitic lifecycle. |
| Fertilization and ookinete | Immunofluorescence microscopy on non-permeabilized parasites confirmed PDI-Trans-GFP expression at the surface of activated male gametes and ookinetes. Live microscopy of mixed blood stages, female gametes and fixed immunofluorescence of sporozoites demonstrated that PDI-Trans-GFP is expressed across the entire parasitic lifecycle. |
| Oocyst | Not tested |
| Sporozoite | Immunofluorescence microscopy on non-permeabilized parasites confirmed PDI-Trans-GFP expression at the surface of activated male gametes and ookinetes. Live microscopy of mixed blood stages, female gametes and fixed immunofluorescence of sporozoites demonstrated that PDI-Trans-GFP is expressed across the entire parasitic lifecycle. |
| Liver stage | Not tested |
| Additional remarks phenotype | Mutant/mutation
The mutant expresses a C-terminal EGFP-tagged version of PDI-Trans
Protein (function)
Protein Disulphide Isomerase (PDI) (EC: 5.3.4.1) is a multifunctional member of the thioredoxin superfamily of redox proteins, characterized by the presence of the βαβαβαββα fold. PDIs typically have three catalytic activities; disulphide isomerase, thiol-disulphide oxidoreductase, and redox-dependent chaperone. PDI homologues have been identified in multiple species, where they are “classically” located in the endoplasmic reticulum (ER) and facilitate the folding and assembly of secretory and membrane proteins within the lumen. In Plasmodium, a small number of proteins have been putatively identified (by sequence homology) as PDI-like molecules in Plasmodium falciparum, vivax, knowlesi, berghei and yoelii. Conclusive demonstration of PDI activity has currently only been demonstrated with PF3D7_0827900/PDI-8, with transcription and translation demonstrated in asexual blood schizonts, gametocytes and sporozoites. Knowledge regarding the process of disulphide bond-dependent protein folding in Plasmodium is scarce.
Previous proteomic analysis of a P. berghei male gamete proteome followed by advanced bioinformatics analysis encompassing a suite of functional and localization-based algorithms identified the expression of PDI-Trans (PBANKA_0820300) in the male gamete, and suggested that the resulting transmembrane protein was potentially located on the surface of the plasma membrane of male gametes.
Phenotype
Phenotype analyses of a mutant lacking expression of PDI-Trans (RMgm-4710) showed the following:
Normal production of gametocytes; normal activation, gamete formation and egress of gametocytes/gametes from host red blood cells. No ookinete formation in vitro. Crossing experiments with mutants defective in either male or female gamete production showed that ΔPDI-Trans female gametes are fertile while ΔPDI-Trans male gamete fertility is strongly reduced.
Strongly reduced oocyst formation (63-96% inhibition) of oocyst formation; the small number of oocysts formed are morphologically normal and form normal sporozoites that are infective to mice.
Analyses of the mutant expressing a C-terminal EGFP-tagged version of PDI-Trans showed:
Immunofluorescence microscopy on non-permeabilized parasites confirmed PDI-Trans-GFP expression at the surface of activated male gametes and ookinetes. Live microscopy of mixed blood stages, female gametes and fixed immunofluorescence of sporozoites demonstrated that PDI-Trans-GFP is expressed across the entire parasitic lifecycle.
Additional information
Other mutants |
*RMgm-4711
