Mutant/mutation
The mutant expresses a mutated form of QC (cyclase dead QC; containing two point mutations in the catalytic site; F103A, Q105A). In addition, it expresses mCherry and luciferase under control of constitutive promoters. The mutated QC gene has been introduced by GIMO transfection in mutant PbΔqc-G (3198cl1; RMgm-5058).

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
The importance of QC cyclase activity to prevent melanization, was analysed using a P. berghei mutant expressing a cyclase-dead QC containing two point mutations in the catalytic site (F103A, Q105A). Similar to QC-null mutants, melanization was observed in this mutant with reduced sg-sporozoite numbers, revealing that cyclase-activity is critical for preventing melanization.

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'.