RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
MutatedGene model (rodent): PBANKA_1429300; Gene model (P.falciparum): PF3D7_1213500; Gene product: integral membrane protein GPR180, putative (GPR180)
Details mutation: The endogenous P. berghei gpr180 gene replaced with the grp180 ortholog of P. vivax (PVX_123365)
PhenotypeNo phenotype has been described
Last modified: 29 April 2022, 14:07
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene mutation
Reference (PubMed-PMID number) Reference 1 (PMID number) : 35404079
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone P. berghei ANKA 2.34
Other information parent lineP. berghei ANKA 2.34 is a cloned, gametocyte producer line of the ANKA strain (PubMed: PMID: 15137943).
The mutant parasite was generated by
Name PI/ResearcherWang PP, Zhu Z
Name Group/DepartmentDepartment of Immunology, College of Basic Medical Sciences
Name InstituteChina Medical University
Name of the mutant parasite
RMgm numberRMgm-5198
Principal namePvGPR180-transgenic parasites
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageNot different from wild type
Gametocyte/GameteNot different from wild type
Fertilization and ookineteNot different from wild type
OocystNot different from wild type
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

In the mutant the endogenous P. berghei gpr180 gene replaced with the gpr180 ortholog of P. vivax (which is C-terminal tagged with 3xHA)

Protein (function)
Heptahelical serpentine receptors are the largest group of membrane receptors responsible for transducing extracellular signals to various downstream effectors. The serpentine receptors coupled to heterotrimeric guanine nucleotide-binding proteins belong to G-protein-coupled receptors (GPCRs) with a salient feature of seven transmembrane domains, each consisting of 25–35 residues. Although there is little conservation in amino acid (aa) sequences across the entire GPCR superfamily, they share similar structures, which are used to classify the GPCRs into six main classes (A – F). Rhodopsin-like Class A is the largest class, accounting for around 90% of GPCRs. Structurally, Rhodopsin-like GPCRs have a GpcrRhopsn4 domain, an eighth helix, and a palmitoylated cysteine at the C-terminal tail. Bioinformatic analysis identified a gpr180-like gene in all Plasmodium species, with the GPCR-like transmembrane domain located in the C terminus, as predicted using the HMMER program. The ~250 aa GPCR domain contains residues that are highly conserved in the Rhodopsin-like GPCR transmembrane domain, which classifies the GPR180 proteins as Class A (Rhodopsin-like) family members of GPCRs

The absence of a phenotype for gametes and ookinetes indicate that PPR180 from P. vivax can functionally complement P. berghei GPR180.

Analysis of a mutant lacking expression of GPR180 (see RMgm-5196) indicates a function during male and female gamete formation (normal (wild type) numbers of gametocytes are produced; male gamete formation ~2fold reduced (as determined by counting exflagellation); female gamete formation reduced by ~ 30%. Crossing experiments indicate a more severe defect in female gametes compared to males. Reduced ookinete formation).

Additional information
Expression of GPR180 in schizonts, gametocytes and ookinetes with highest expression in gametocytes (see mutant RMgm-5197 that expresses HA-tagged GPR180). Evidence is presented that gametocyte activation stimulates GPR180 redistribution from the cytoplasm to the plasma membrane.

Other mutants

  Mutated: Mutant parasite with a mutated gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1429300
Gene Model P. falciparum ortholog PF3D7_1213500
Gene productintegral membrane protein GPR180, putative
Gene product: Alternative nameGPR180
Details of the genetic modification
Short description of the mutationThe endogenous P. berghei gpr180 gene replaced with the grp180 ortholog of P. vivax (PVX_123365)
Inducable system usedNo
Short description of the conditional mutagenesisNot available
Additional remarks inducable system
Type of plasmid/construct(Linear) plasmid double cross-over
PlasmoGEM (Sanger) construct/vector usedNo
Modified PlasmoGEM construct/vector usedNo
Plasmid/construct map
Plasmid/construct sequence
Restriction sites to linearize plasmid
Selectable marker used to select the mutant parasitetgdhfr
Promoter of the selectable markereef1a
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationTo replace the endogenous pbgpr180 with the pvgpr180 ORF, an 1157 bp fragment containing the 5’UTR of pbgpr180 (nt: 21157 bp – 21 bp), the 2485 bp pvgpr180 (PlasmoDB ID: PVX_123365) ORF, and the 871 bp 3R were amplified and cloned into HindIII/ApaI, ApaI/SacII, and XhoI/NotI sites of the pL0034 vector, respectively, to generate pL0034-PvGPR180-R. The pL0034-PvGPR180-R plasmid was linearized with BglI and NotI.
Additional remarks selection procedure
Primer information: Primers used for amplification of the target sequences  Click to view information
Primer information: Primers used for amplification of the target sequences  Click to hide information
Sequence Primer 1
Additional information primer 1
Sequence Primer 2
Additional information primer 2
Sequence Primer 3
Additional information primer 3
Sequence Primer 4
Additional information primer 4
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6