Sporozoite | Reduced numbers of salivary gland sporozoites. Salivary gland sporozoites show wild-type morphology, motility and infectivity to hepatocytes in vitro. Light-microscopy analysis of oocysts between day 14 and 21 showed the presence of aberrant, enlarged, melanized sporozoites, either still inside or during release from oocysts.
On day 21 p.i. non-motile, enlarged sporozoites covered by melanin were found in the hemocoel, often in clusters or attached to salivary glands. In contrast, most QC-null sporozoites inside salivary glands had a WT-like morphology, i.e. long and slender, without signs of melanization. |
Additional remarks phenotype | Mutant/mutation
The mutant lacks expression of QC and expresses mCherry and luciferase under control of constitutive promoters. In this QC-GIMO line the qc open reading frame is replaced with a hdhfr-yfcu selectable marker cassette.
Protein (function)
N-terminal modification of glutamine or glutamic acid residues to pyroglutamic acid (pGlu; 5-oxo-L-46 proline) is a posttranslational modification (PTM), catalyzed by glutaminyl cyclases (QCs) found in eukaryotes and prokaryotes. Two evolutionary unrelated classes exist; mammalian QCs and QCs of bacteria, plants and parasites, which share no sequence homology, supporting a different evolutionary origin. Mammalian cells can express two forms, the secreted glutaminyl‐peptide cyclotransferase (QPCT) or its iso-enzyme (QPCTL), localized in the Golgi complex. pGlu is implicated in maturation and stabilization of mammalian proteins such has neuropeptides and cytokines. QC activity has been associated in humans with pathological processes such as amyloidotic diseases and QPCTL is critical for pGLu formation on CD47, facilitating myeloid immune evasion.
A single gene encoding a glutaminyl cyclase (QC), named glutaminyl-peptide cyclotransferase, has been identified by electronic annotation in all sequenced Plasmodium genomes. Plasmodium QCs share 70-76% sequence similarity and 50-54% identity and contain a transmembrane domain. QC of the human malaria parasite P. falciparum (PfQC) shows 21-27% identity to QCs of various 64 bacteria and the plant Carica papaya (CpQC). All 9 amino acids of the catalytic site of bacterial and plant QCs are conserved in Plasmodium QC.
Phenotype
Normal numbers of oocysts are produced. On day 10 after infection melanized oocysts are present in 55-65% of QC-null infected mosquitoes (1-85 melanized oocysts/mosquito), while such oocysts were absent in WT-infected mosquitoes.
Reduced numbers of salivary gland sporozoites. Salivary gland sporozoites show wild-type morphology, motility and infectivity to hepatocytes in vitro. Light-microscopy analysis of oocysts between day 14 and 21 showed the presence of aberrant, enlarged, melanized sporozoites, either still inside or during release from oocysts.
On day 21 p.i. non-motile, enlarged sporozoites covered by melanin were found in the hemocoel, often in clusters or attached to salivary glands. In contrast, most QC-null sporozoites inside salivary glands had a WT-like morphology, i.e. long and slender, without signs of melanization.
Wild type development of liver stages in vitro after infection of hepatocytes with salivary gland sporozoites.
Additional information
Published expression data indicate that QC is expressed in gametocytes, ookinetes, oocysts, and sporozoites. QC expression in mosquito/transmission stages was confirmed by analyzing a transgenic rodent malaria parasite (P. berghei; Pbqc::cmyc), expressing a cmyc-tagged QC (RMgm-5059).
From the Abstract:
'We show that Plasmodium sporozoites of QC-null mutants are recognized by the mosquito immune system and melanized when they reach the hemocoel. Sporozoite numbers in salivary glands are also reduced in mosquitoes infected with QC-null or QC catalytically-dead mutants. This phenotype can be rescued by genetic complementation or by disrupting mosquito hemocytes or melanization immune responses. Mutation of a single QC-target glutamine of the major sporozoite surface protein (CSP) also results in immune recognition of sporozoites. These findings reveal QC-mediated post-translational modification of surface proteins as a major mechanism of mosquito immune evasion by Plasmodium sporozoites'.
Other mutants |