Summary

RMgm-1340
Malaria parasiteP. berghei
Genotype
DisruptedGene model (rodent): PBANKA_1034300; Gene model (P.falciparum): PF3D7_1408200; Gene product: AP2 domain transcription factor AP2-G2, putative (AP2-G2; ApiAP2)
Phenotype Gametocyte/Gamete; Fertilization and ookinete; Oocyst;
Last modified: 25 October 2015, 12:54
  *RMgm-1340
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) : 26417110
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/ResearcherYuda M; Kato T
Name Group/DepartmentDepartment of Medical Zoology
Name InstituteMie University School of Medicine
CityMie
CountryJapan
Name of the mutant parasite
RMgm numberRMgm-1340
Principal nameAP2-G2(−)
Alternative name
Standardized name
Is the mutant parasite cloned after genetic modificationYes
Phenotype
Asexual blood stageNot different from wild type
Gametocyte/GameteNo mature male or female gametocytes are formed. No exflagellation. Evidence is presented that sexually committed parasites (trophozoites) are formed within red blood cells but that these parasites abort development before morphological features of sexual differentiation are visible at the light-microscopic level.
Fertilization and ookineteNo mature male or female gametocytes are formed. No exflagellation. No oocysts are formed.
OocystNo mature male or female gametocytes are formed. No exflagellation. No oocysts are formed.
SporozoiteNot tested
Liver stageNot tested
Additional remarks phenotype

Mutant/mutation
The mutant lacks expression of AP2-G2

Protein (function)
Apetala 2 (AP2)-family proteins are transcription factors that have DNA-binding domains of 60 amino acids called AP2 domains. Recently, AP2 genes have been found in the genomes of Plasmodium parasites. In P. falciparum 27 AP2-family genes have been identified. Among these genes, 26 are conserved in the other Plasmodium species whose entire genomes have been sequenced. Each member of this family has 1 to 4 AP2 domains, and the amino acid sequences of these domains are highly conserved among Plasmodium orthologs.

Phenotype
No mature male or female gametocytes are formed. No exflagellation. Evidence is presented that sexually committed parasites (trophozoites) are formed within red blood cells but that these parasites abort development before morphological features of sexual differentiation are visible at the light-microscopic level.

Evidence is presented that the lack of AP2-G2 expression did not abrogate sexual commitment or sex determination but did cause marked developmental defects in the gametocytes, together with a marked reduction of sex-specific gene expression.

Evidence is presented that AP2-G2 is a transcriptional repressor in sexually committed trophozoites. Evidence is presented that most of these target genes of AP2-G2 binding are required for asexual proliferation of the parasites in the blood, suggesting that AP2-G2 blocks the program of asexual replication to promote conversion to the sexual stage. These 'asexual' genes were upregulated in the parasites lacking expression of AP2-G2

Additional information
In analyses using mutant P. berghei parasites expressing GFP-tagged AP2-G2 (AP2-G2::GFP parasites; RMgm-1342), it was found that AP2-G2 was specifically expressed in female and male gametocytes. The GFP signal was solely observed in the nucleus, suggesting that AP2-G2 acts as a transcription factor in gametocytes. In a temporal profiling analysis of AP2-G2 expression using synchronized blood cultures, expression was first observed from 16 h after erythrocyte invasion (hpi) until development into mature male or female gametocytes. The timing of expression correlated well with the onset of morphological divergence from the asexual stage (16–18 hpi), strongly suggesting that this transcription factor is involved in gametocyte-specific gene expression.

Other mutants
See link for other AP2-G2 mutants


  Disrupted: Mutant parasite with a disrupted gene
Details of the target gene
Gene Model of Rodent Parasite PBANKA_1034300
Gene Model P. falciparum ortholog PF3D7_1408200
Gene productAP2 domain transcription factor AP2-G2, putative
Gene product: Alternative nameAP2-G2; ApiAP2
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 markerpbdhfr
Selection (positive) procedurepyrimethamine
Selection (negative) procedureNo
Additional remarks genetic modification
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 15´-ACCACGATAAAGGAGGCATGTAAC-3´
Additional information primer 1AP2-G2-1
Sequence Primer 25´-CTCATCTACAAGCATCgtcgacTCATCGTCTCCTTTCCTC-3´
Additional information primer 2AP2-G2-2
Sequence Primer 35´-CCTTCAATTTCGgatccactagGTAATGTTGAAAGCGACAG-3´
Additional information primer 3AP2-G2-3
Sequence Primer 45´-GTTGTATTATCAACTTGAGCAGTTTC-3´
Additional information primer 4AP2-G2-4
Sequence Primer 5
Additional information primer 5
Sequence Primer 6
Additional information primer 6