IsoQC (QPCTL) knock-out mice suggest differential substrate conversion by glutaminyl cyclase isoenzymes

Abstract

Secretory peptides and proteins are frequently modified by pyroglutamic acid (pE, pGlu) at their N-terminus. This modification is catalyzed by the glutaminyl cyclases QC and isoQC. Here, we decipher the roles of the isoenzymes by characterization of IsoQC-/- mice. These mice show a significant reduction of glutaminyl cyclase activity in brain and peripheral tissue, suggesting ubiquitous expression of the isoQC enzyme. An assay of substrate conversion in vivo reveals impaired generation of the pGlu-modified C-C chemokine ligand 2 (CCL2, MCP-1) in isoQC-/- mice. The pGlu-formation was also impaired in primary neurons, which express significant levels of QC. Interestingly, however, the formation of the neuropeptide hormone thyrotropin-releasing hormone (TRH), assessed by immunohistochemistry and hormonal analysis of hypothalamic-pituitary-thyroid axis, was not affected in isoQC-/-, which contrasts to QC-/-. Thus, the results reveal differential functions of isoQC and QC in the formation of the pGlu-peptides CCL2 and TRH. Substrates requiring extensive prohormone processing in secretory granules, such as TRH, are primarily converted by QC. In contrast, protein substrates such as CCL2 appear to be primarily converted by isoQC. The results provide a new example, how subtle differences in subcellular localization of enzymes and substrate precursor maturation might influence pGlu-product formation.

Immunohistochemical Evidence from APP-Transgenic Mice for Glutaminyl Cyclase as Drug Target to Diminish pE-Abeta Formation

“…In addition to pathogenic N-truncated Abeta peptides, there are also physiological QC substrates, such as neuropeptides and peptide hormones [ 39 , 40 ], that could be potentially affected by QC inhibition. However, in experimental animals with genetic QC ablation or pharmacological QC inhibition [ 41 , 42 , 43 ], it was found that the hypothalamic-gonadotropic and hypothalamic-thyroid axes, which are regulated by the QC substrates gonadoliberin (GnRH) and thyroliberin (TRH), are only marginally affected in presence of strong QC inhibition. Furthermore, in two clinical trials (see below), no effects on these hormone axes or other side effects with relation to QC substrates were observed, although a QC target occupancy in the brain of about 90% was reached under treatment.…”

Identification of Glutaminyl Cyclase Genes Involved in Pyroglutamate Modification of Fungal Lignocellulolytic Enzymes

“…A second QC in mammals is retained within the Golgi complex (isoQC) (15, 39). Differential cellular distribution of QC and isoQC may indicate a preference for different substrates in the secretory pathway and thus distinct physiological roles.…”

“…Differential cellular distribution of QC and isoQC may indicate a preference for different substrates in the secretory pathway and thus distinct physiological roles. This has been demonstrated in monocyte chemoattractant proteins (MCPs) versus thyrotropin-releasing hormone (TRH) where only one QC is involved in the pGlu formation for each protein (39, 40). In N. crassa , both QC-1 and QC-2 localized primarily to the ER, where proteins with a liberated N-terminal Q or E would be cyclized to pGlu.…”

Purification of recombinant Aβ(1‐42) and pGlu‐Aβ(3‐42) using preparative SDS‐PAGE

“…Mouse primary neurons were isolated as described by Becker et al. (2015) . The cultivated cells were treated with freshly dissolved Aβ(1‐42) and pGlu‐Aβ(3‐42) at a final concentration of 5 μM for 69 h. Afterwards, MTT (3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl tetrazolium bromide) has been added to the cell medium.…”