Mutant/mutation
The mutant lacks expression of DPAP3 and expresses the GFP-Luciferase fusion protein under control of the (schizont specific) ama-1 promoter.

Protein (function)
The P. falciparum and P. berghei genomes each contain three dipeptidyl aminopeptidase (dpap) homologs. P. falciparum DPAP1 and DPAP3 are critical for asexual growth. The P. falciparum dpap2 gene could be deleted. Parasites lacking expression of (the gametocyte-specific?) DPAP2 produced normal gametocytes.

DPAP3 of P. falciparum has been implicated in egress of merozoites from the host erythrocyte.

Evidence has been presented that the P. falciparum enzyme dipeptidyl aminopeptidase 1 (DPAP1; an ortholog of the lysosomal exopeptidase cathepsin C) was located in the food/digestive vacuole and to possesses hydrolytic activity against dipeptide substrates, suggesting that DPAP1 is involved in the generation of dipeptides from hemoglobin-derived oligopeptides. The dpap1 gene could not be deleted from the P. falciparum genome, suggesting an essential role during blood stage growth/multiplication.

In P. berghei all three dpap genes can be deleted from the genome (see below).

Phenotype
Blood stages showed a normal growth/development (blood stages produced normal levels of hemozoin). See also 'additional information' on the reduction of experimental cerebral malaria complications (ECM) in mice infected with this mutant

Additional information
See below for genotyping:

This mutant is also used in the study: Capuccini B et al (2016). Sci Rep. 2016 Dec 19;6:39258 (PMID: 27991544). The following was found:
'In order to determine whether the changes in microglia revealed in the microarray were generated only in an infection that gave rise to ECM, we compared responses of microglia obtained at day 7 from ECM-causing PbA infection with that of a mutant of PbA, which does not cause ECM. For this we used a gene-deletion mutant for the dipeptidyl peptidase 3 (DPAP3), Δdpap3. Infection with 105 or 104 iRBC of PbA Δdpap3 resulted in a gradually increasing parasitemia and approximately 85% to 100% survival of mice up to 20 days of infection, with low or no clinical signs of ECM. For the first 5 to 7 days of infection the parasitemias were similar to those of wt PbA.
There was no significant increase in the numbers of microglia recovered at day 7 from the PbA Δdpap3 infection in contrast to the 3-fold increase observed in microglia from the wt PbA infection. We performed quantitative RT-qPCR on microglia isolated from both infections for the genes, Oasl1, IRF7, H2-Q8, Cxcl10 and Ccl5. Only microglia from wt PbA infections causing ECM show significant fold increases in expression of any of these genes (Fig. 4e). These data suggest that activation of microglia may be feature of ECM'.

Genotyping:

Generation of the P. berghei mutants ∆dpap1 (RMgm-810), ∆dpap2 (RMgm-811) and ∆dpap3 (RMgm-812).
(primer sequences can either be found in Lin et al. (2015). J. Exp. Med. or obtained from the Leiden Malaria Research Group)
(A) Schematic representation of the gene-deletion constructs targeting the ORF of genes expressing dipeptidyl peptidases 1-3 (dpap1-3) by double cross-over homologous recombination and the wt gene loci before and after disruption. The constructs contain a drug-selectable marker cassette (SM; black box) and gene target regions (hatched boxes). Primer positions (arrows) for diagnostic PCRs are shown (see Table S4 for primer sequences and expected product sizes). (B) Diagnostic PCR (left, center) and RT-PCR (right) analysis confirm correct disruption of dpap1 in ∆dpap1-a. For diagnostic PCRs, the following primers were used: 5’ in, RS672/RS32; 3’ in, RS110/RS673; SM (tgdhfr/ts), RS404/RS405; dpap1 ORF, RS582/RS583. For RT-PCR the following primers were used: tub (tubulin), RS782/RS783 and dpap1, RS582/RS583. (C) Diagnostic PCR (left) and Southern analysis of pulsed field gel-separated (center) confirm correct disruption of dpap1 in ∆dpap1-b. Northern analysis of blood-stage mRNA (right) confirms the absence of dpap1 transcripts in the ∆dpap1-a mutant. The following primers were used for diagnostic PCRs: 5' integration (5’ in), L6204/L4770; 3' integration (3’ in), L4771/L6205; SM (hdfhr::yfcu), L4698/L4699; dpap1 ORF, L6206/L6207. For Southern analysis, separated chromosomes were hybridized using an hdhfr probe that recognizes the construct integrated into the dpap1 locus on chromosome 9. Northern blot was hybridized using a PCR probe recognizing the dpap1 ORF (primers L6206/L6207). As a loading control, hybridization was performed with oligonucleotide probe L644R that recognizes the large subunit rRNA. (D) Diagnostic PCR (left) and Southern analysis of pulsed field gel-separated chromosomes (center) confirms correct disruption of dpap2 in mutant ∆dpap2. Northern analysis of blood-stage mRNA (right) confirms the absence of dpap2 transcripts in the ∆dpap2 mutant. The following primers were used for diagnostic PCRs: 5’ in, L6935/L4770; 3’ in, L4771/L6936; SM (hdfhr::yfcu), L4698/L4699; dpap2 ORF, L6937/L6938. Separated chromosomes were hybridized using a 3’UTR pbdhfr probe that recognizes the construct integrated into dpap2 on chromosome 14, the endogenous dhfr/ts on chromosome 7 and the GFP-luciferase reporter cassette in the 230p locus on chromosome 3. Northern blot was hybridized using a PCR probe recognizing the dpap2 ORF (primers L6937/L6938) and with an oligonucleotide probe L644R recognizing the large subunit rRNA (as loading control). (E) Diagnostic PCR (left) and Southern analysis of pulsed field gelseparated chromosomes (center) confirm correct disruption of dpap3 in ∆dpap3-a and ∆dpap3-b. Northern analysis of blood-stage mRNA (right) confirms the absence of dpap3 transcripts. The following primers were used for diagnostic PCRs: 5’ in, L6941/L4770; 3’ in, L4771/L6942; SM (hdfhr::yfcu), L4698/L4699; dpap3 ORF, L6943/L6944. Separated chromosomes were hybridized using a 3’UTR pbdhfr probe that recognizes the construct integrated into dpap3 on chromosome 10, the endogenous dhfr/ts on chromosome 7 and the GFP-luciferase reporter cassette in the 230p locus on chromosome 3. Northern blot was hybridized using a PCR probe recognizing the dpap3 ORF (primers L6943/L6944) and with an oligonucleotide probe L644R recognizing the large subunit rRNA (as loading control). See Table S4 for primers used for generation of probes.

Other mutants
RMgm-810: mutant lacking expression of DPAP1
RMgm-811: mutant lacking expression of DPAP2