RMgmDB - Rodent Malaria genetically modified Parasites


Malaria parasiteP. berghei
DisruptedGene model (rodent): PBANKA_1205000; Gene model (P.falciparum): PF3D7_1006800; Gene product: single-strand telomeric DNA-binding protein GBP2, putative (GBP2, G-strand binding protein 2)
Phenotype Asexual bloodstage; Gametocyte/Gamete;
Last modified: 26 October 2021, 13:37
Successful modificationThe parasite was generated by the genetic modification
The mutant contains the following genetic modification(s) Gene disruption
Reference (PubMed-PMID number) Reference 1 (PMID number) : 31958569
Reference 2 (PMID number) : 34604117
MR4 number
Parent parasite used to introduce the genetic modification
Rodent Malaria ParasiteP. berghei
Parent strain/lineP. berghei ANKA
Name parent line/clone Not applicable
Other information parent line
The mutant parasite was generated by
Name PI/ResearcherNiikura M, Kobayashi F
Name Group/DepartmentDepartment of Environmental Science, School of Life and Environmental Science
Name InstituteAzabu University
Name of the mutant parasite
RMgm numberRMgm-4707
Principal nameΔgbp2
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Asexual blood stageIn the first paper: Reduced growth/multiplication of asexual blood stages. Blood stage infections induce experimental cerebral malaria (ECM) in C57Bl6 mice.
In the second paper: normal (wild type) growth of asexual blood stages.
Gametocyte/Gamete(Strongly) reduced numbers of male and female gametocytes
Fertilization and ookineteNot tested
OocystNot tested
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

The mutant lacks expression of GBP2

Protein (function)
G-strand binding protein 2 (GBP2) is a Ser/Arg-rich (SR) protein involved in mRNA surveillance and nuclear mRNA quality control in yeast. PfGBP2 (PF3D7_1006800) has homology to yeast GBP2 and is highly expressed during the blood stages of malaria parasites. GBP2 of Plasmodium spp. contains two RNA recognition motif (RRM) domains (RRM1 and RRM2) and is conserved among eukaryotes. RRM1 and RRM2 of yeast GBP2 recognize RNA. RRM2 of yeast GBP2 contains a WQxLKD motif, which is directly involved in RNA recognition. Plasmodium RRM2 contains a WKELKD motif, which is similar to the WQxLKD motif. In contrast, RRM3 (the C-terminal RRM domain of yeast) was absent in Plasmodium spp.

In the first paper: Reduced growth/multiplication of asexual blood stages. Blood stage infections induce experimental cerebral malaria (ECM) in C57Bl6 mice.
In the second paper: normal (wild type) growth of asexual blood stages..
(Strongly) reduced numbers of male and female gametocytes.

Additional information
To investigate the cellular localization of GBP2, we generated transgenic parasites expressing GBP2-mCherry fusion protein (GBP2::mCherry; RMgm-5086). The mCherry tag was introduced at the C-terminus of endogenous GBP2. Gbp2::mCherry expression was controlled by the endogenous gbp2 native promoters.
To examine the cellular localization of GBP2, we performed live-cell fluorescence imaging of cultured GBP2::mCherry schizonts and infected erythrocytes obtained from mice at 6 h (ring form), 12 h (trophozoite) and 18 h (late trophozoite) post-inoculation with GBP2::mCherry schizonts The mCherry signal was distributed throughout GBP2::mCherry parasite cells at all development stages. GBP2 localized to both the nucleus and cytoplasm of malaria parasites.  
To investigate which proteins interact with GBP2, we performed protein IP using anti-mCherry beads and identified the proteins bound to GBP2 by MS. The results of IP-MS  of GBP2::mCherry imply that GBP2 interacts with ALBA4, DOZI and CITH. DOZI and CITH are required for zygote development but not for gametocytogenesis.
To identify RNAs bound by GBP2 ,we performed RIPseq on mature schizonts- and gametocytes-lysates using antimCherry beads and sequenced the RNAs bound to GBP2 Our RIP-seq results revealed that GBP2 bound to mRNAs encoding proteins that are essential for asexual development; however, the parasitemia course was comparable in mice infected with Δgbp2 versus control parasites. These findings imply that a factor other than GBP2 is involved in the export of these mRNAs.

Other mutants

  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1205000
Gene Model P. falciparum ortholog PF3D7_1006800
Gene productsingle-strand telomeric DNA-binding protein GBP2, putative
Gene product: Alternative nameGBP2, G-strand binding protein 2
Details of the genetic modification
Inducable system usedNo
Additional remarks inducable system
Type of plasmid/construct used(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
Partial or complete disruption of the geneComplete
Additional remarks partial/complete disruption
Selectable marker used to select the mutant parasitehdhfr
Promoter of the selectable markerunknown
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modificationThe gene-targeting vector for GBP2, gbp2 (PBANKA_120500), was prepared by PCR. Briefly, the 5'and 3' flanking regions of the gbp2 open reading frame (ORF) were amplified by PCR. The PCR products were annealed to either side of the red fluorescent protein gene (mCherry)-hdhfr-expressing cassette and amplified by PCR using gene-specific primers
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