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

  • Andreas Becker , Rico Eichentopf , Reinhard Sedlmeier , Alexander Waniek , Holger Cynis , Birgit Koch , Anett Stephan , Christoph Bäuscher , Stephanie Kohlmann , Torsten Hoffmann , Astrid Kehlen , Sabine Berg , Ernst-Joachim Freyse , Alexander Osmand , Anne-Christine Plank , Steffen Roßner , Stephan von Hörsten , Sigrid Graubner , Hans-Ulrich Demuth und Stephan Schilling EMAIL logo
Veröffentlicht/Copyright: 19. August 2015
Biological Chemistry
Aus der Zeitschrift Biological Chemistry Band 397 Heft 1

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.

Keywords: 5-oxo-L-proline; posttranslational modification; pyroglutamate; TRH
Corresponding author: Stephan Schilling, Probiodrug AG, Weinbergweg 22, D-06120 Halle/Saale, Germany, e-mail:
aAndreas Becker and Rico Eichentopf: These authors contributed equally to this work.bPresent address: Fraunhofer Institute for Cell Therapy and Immunology

Acknowledgments

The authors thank D. Friedrich, E. Scheel and H.-H. Ludwig for their excellent technical assistance as well as K. Arendt for critically reading of the manuscript. This work was financially supported by the Investitionsbank Sachsen-Anhalt, grant# 1004/00082 to Probiodrug AG.

References

Alexandru, A., Jagla, W., Graubner, S., Becker, A., Bäuscher, C., Kohlmann, S., Sedlmeier, R., Raber, K., Cynis, H., Rönicke, R., et al. (2011). Selective hippocampal neurodegeneration in transgenic mice expressing small amounts of truncated Abeta is induced by pyroglutamate-Aβ formation. J. Neurosci. 31, 12790–12801.10.1523/JNEUROSCI.1794-11.2011Suche in Google Scholar PubMed PubMed Central

Augustin, M., Sedlmeier, R., Peters, T., Huffstadt, U., Kochmann, E., Simon, D., Schöniger, M., Garke-Mayerthaler, S., Laufs, J., Mayhaus, M., et al. (2005). Efficient and fast targeted production of murine models based on ENU mutagenesis. Mamm. Genome 16, 405–413.10.1007/s00335-004-3028-2Suche in Google Scholar PubMed

Böckers, T., Kreutz, M., and Pohl, T. (1995). Glutaminyl-cyclase expression in the bovine/porcine hypothalamus and pituitary. J. Neuroendocrinol. 7, 445–453.10.1111/j.1365-2826.1995.tb00780.xSuche in Google Scholar PubMed

Cruz, I. and Nillni, E. (1996). Intracellular sites of prothyrotropin- releasing hormone processing. J. Biol. Chem. 271, 22736–22745.10.1074/jbc.271.37.22736Suche in Google Scholar PubMed

Cynis, H., Schilling, S., Bodnár, M., Hoffmann, T., Heiser, U., Saido, T., and Demuth, H. (2006). Inhibition of glutaminyl cyclase alters pyroglutamate formation in mammalian cells. Biochim. Biophys. Acta 1764, 1618–1625.10.1016/j.bbapap.2006.08.003Suche in Google Scholar PubMed

Cynis, H., Rahfeld, J., Stephan, A., Kehlen, A., Koch, B., Wermann, M., Demuth, H., and Schilling, S. (2008). Isolation of an isoenzyme of human glutaminyl cyclase: retention in the Golgi complex suggests involvement in the protein maturation machinery. J. Mol. Biol. 379, 966–980.10.1016/j.jmb.2008.03.078Suche in Google Scholar PubMed

Cynis, H., Hoffmann, T., Friedrich, D., Kehlen, A., Gans, K., Kleinschmidt, M., Rahfeld, J., Wolf, R., Wermann, M., Stephan, A., et al. (2011). The isoenzyme of glutaminyl cyclase is an important regulator of monocyte infiltration under inflammatory conditions. EMBO Mol. Med. 3, 545–558.10.1002/emmm.201100158Suche in Google Scholar PubMed PubMed Central

De Kimpe, L., Bennis, A., Zwart, R., van Haastert, E., Hoozemans, J., and Scheper, W. (2012). Disturbed Ca2+ homeostasis increases glutaminyl cyclase expression; connecting two early pathogenic events in Alzheimer’s disease in vitro. PLoS One 7, e44674.10.1371/journal.pone.0044674Suche in Google Scholar PubMed PubMed Central

Gong, J. and Clark-Lewis, I. (1995). Antagonists of monocyte chemoattractant protein 1 identified by modification of functionally critical NH2-terminal residues. J. Exp. Med. 181, 631–640.10.1084/jem.181.2.631Suche in Google Scholar PubMed PubMed Central

Goren, H., Bauce, L., and Vale, W. (1977). Forces and structural limitations of binding of thyrotrophin-releasing factor to the thyrotrophin-releasing receptor: the pyroglutamic acid moiety. Mol. Pharmacol. 13, 606–614.Suche in Google Scholar

Hartlage-Rübsamen, M., Staffa, K., Waniek, A., Wermann, M., Hoffmann, T., Cynis, H., Schilling, S., Demuth, H., and Rossner, S. (2009). Developmental expression and subcellular localization of glutaminyl cyclase in mouse brain. Int. J. Dev. Neurosci. 27, 825–835.10.1016/j.ijdevneu.2009.08.007Suche in Google Scholar PubMed

Hartlage-Rübsamen, M., Morawski, M., Waniek, A., Jäger, C., Zeitschel, U., Koch, B., Cynis, H., Schilling, S., Schliebs, R., Demuth, H., et al. (2011). Glutaminyl cyclase contributes to the formation of focal and diffuse pyroglutamate (pGlu)-Aβ deposits in hippocampus via distinct cellular mechanisms. Acta Neuropathol. 121, 705–719.10.1007/s00401-011-0806-2Suche in Google Scholar

Höfling, C., Indrischek, H., Höpcke, T., Waniek, A., Cynis, H., Koch, B., Schilling, S., Morawski, M., Demuth, H.U., Roßner, S., et al. (2014). Mouse strain and brain region-specific expression of the glutaminyl cyclases QC and isoQC. Int. J. Dev. Neurosci. 36, 64-73.10.1016/j.ijdevneu.2014.05.008Suche in Google Scholar

Huang, K., Liu, Y., Cheng, W., Ko, T., and Wang, A. (2005). Crystal structures of human glutaminyl cyclase, an enzyme responsible for protein N-terminal pyroglutamate formation. Proc. Natl. Acad. Sci. USA 102, 13117–13122.10.1073/pnas.0504184102Suche in Google Scholar

Huang, K., Liaw, S., Huang, W., Chia, C., Lo, Y., Chen, Y., and Wang, A. (2011). Structures of human Golgi-resident glutaminyl cyclase and its complexes with inhibitors reveal a large loop movement upon inhibitor binding. J. Biol. Chem. 286, 12439–12449.10.1074/jbc.M110.208595Suche in Google Scholar

Iwatsubo, T., Saido, T., Mann, D., Lee, V., and Trojanowski, J. (1996). Full-length amyloid-beta (1-42(43)) and amino-terminally modified and truncated amyloid-β 42(43) deposit in diffuse plaques. Am. J. Pathol. 149, 1823–1830.Suche in Google Scholar

Jawhar, S., Wirths, O., and Bayer, T. (2011a). Pyroglutamate amyloid-β (Aβ): a hatchet man in Alzheimer disease. J. Biol. Chem 286, 38825–38832.10.1074/jbc.R111.288308Suche in Google Scholar

Jawhar, S., Wirths, O., Schilling, S., Graubner, S., Demuth, H., and Bayer, T. (2011b). Overexpression of glutaminyl cyclase, the enzyme responsible for pyroglutamate Aβ formation, induces behavioral deficits, and glutaminyl cyclase knock-out rescues the behavioral phenotype in 5XFAD mice. J. Biol. Chem 286, 4454–4460.10.1074/jbc.M110.185819Suche in Google Scholar

Li, Q., Liu, Z., Monroe, H., and Culiat, C. (2002). Integrated platform for detection of DNA sequence variants using capillary array electrophoresis. Electrophoresis 23, 1499–1511.10.1002/1522-2683(200205)23:10<1499::AID-ELPS1499>3.0.CO;2-XSuche in Google Scholar

Mason, A., Hayflick, J., Zoeller, R., Young, W., Phillips, H., Nikolics, K., and Seeburg, P. (1986). A deletion truncating the gonadotropin-releasing hormone gene is responsible for hypogonadism in the hpg mouse. Science 234, 1366–1371.10.1126/science.3024317Suche in Google Scholar

Morawski, M., Hartlage-Rübsamen, M., Jäger, C., Waniek, A., Schilling, S., Schwab, C., McGeer, P., Arendt, T., Demuth, H., and Roßner, S. (2010). Distinct glutaminyl cyclase expression in Edinger-Westphal nucleus, locus coeruleus and nucleus basalis Meynert contributes to pGlu-Abeta pathology in Alzheimer’s disease. Acta Neuropathol. 120, 195–207.10.1007/s00401-010-0685-ySuche in Google Scholar

Nillni, E. (1999). Neuroregulation of ProTRH biosynthesis and processing. Endocrine 10, 185–199.10.1007/BF02738618Suche in Google Scholar

Nillni, E. and Sevarino, K. (1999). The biology of pro-thyrotropin-releasing hormone-derived peptides. Endocr. Rev. 20, 599–648.Suche in Google Scholar

Osmand, A., Berthelier, V., and Wetzel, R. (2006). Imaging polyglutamine deposits in brain tissue. Methods Enzymol. 412, 106–122.10.1016/S0076-6879(06)12008-XSuche in Google Scholar

Saido, T., Iwatsubo, T., Mann, D., Shimada, H., Ihara, Y., and Kawashima, S. (1995). Dominant and differential deposition of distinct beta-amyloid peptide species, Aβ N3(pE), in senile plaques. Neuron 14, 457–466.10.1016/0896-6273(95)90301-1Suche in Google Scholar

Saul, A., Lashley, T., Revesz, T., Holton, J., Ghiso, J., Coomaraswamy, J., and Wirths, O. (2013). Abundant pyroglutamate-modified ABri and ADan peptides in extracellular and vascular amyloid deposits in familial British and Danish dementias. Neurobiol. Aging 34, 1416–1425.10.1016/j.neurobiolaging.2012.11.014Suche in Google Scholar PubMed PubMed Central

Schilling, S., Lindner, C., Koch, B., Wermann, M., Rahfeld, J., Bohlen, A., Rudolph, T., Reuter, G., and Demuth, H. (2007). Isolation and characterization of glutaminyl cyclases from Drosophila: evidence for enzyme forms with different subcellular localization. Biochemistry 46, 10921–10930.10.1021/bi701043xSuche in Google Scholar PubMed

Schilling, S., Appl, T., Hoffmann, T., Cynis, H., Schulz, K., Jagla, W., Friedrich, D., Wermann, M., Buchholz, M., Heiser, U., et al. (2008a). Inhibition of glutaminyl cyclase prevents pGlu-Aβ formation after intracortical/hippocampal microinjection in vivo/in situ. J. Neurochem 106, 1225–1236.10.1111/j.1471-4159.2008.05471.xSuche in Google Scholar PubMed

Schilling, S., Zeitschel, U., Hoffmann, T., Heiser, U., Francke, M., Kehlen, A., Holzer, M., Hutter-Paier, B., Prokesch, M., Windisch, M., et al. (2008b). Glutaminyl cyclase inhibition attenuates pyroglutamate Aβ and Alzheimer’s disease-like pathology. Nat. Med. 14, 1106–1111.10.1038/nm.1872Suche in Google Scholar PubMed

Schilling, S., Kohlmann, S., Bäuscher, C., Sedlmeier, R., Koch, B., Eichentopf, R., Becker, A., Cynis, H., Hoffmann, T., Berg, S., et al. (2011). Glutaminyl cyclase knock-out mice exhibit slight hypothyroidism but no hypogonadism: implications for enzyme function and drug development. J. Biol. Chem 286, 14199–14208.10.1074/jbc.M111.229385Suche in Google Scholar PubMed PubMed Central

Sealfon, S., Weinstein, H., and Millar, R. (1997). Molecular mechanisms of ligand interaction with the gonadotropin-releasing hormone receptor. Endocr. Rev. 18, 180–205.10.1210/edrv.18.2.0295Suche in Google Scholar PubMed

Seifert, F., Schulz, K., Koch, B., Manhart, S., Demuth, H., and Schilling, S. (2009). Glutaminyl cyclases display significant catalytic proficiency for glutamyl substrates. Biochemistry 48, 11831–11833.10.1021/bi9018835Suche in Google Scholar PubMed

Stephan, A., Wermann, M., Bohlen, A. von, Koch, B., Cynis, H., Demuth, H., and Schilling, S. (2009). Mammalian glutaminyl cyclases and their isoenzymes have identical enzymatic characteristics. FEBS J. 276, 6522–6536.10.1111/j.1742-4658.2009.07337.xSuche in Google Scholar PubMed

Yamada, M., Saga, Y., Shibusawa, N., Hirato, J., Murakami, M., Iwasaki, T., Hashimoto, K., Satoh, T., Wakabayashi, K., Taketo, M., et al. (1997). Tertiary hypothyroidism and hyperglycemia in mice with targeted disruption of the thyrotropin-releasing hormone gene. Proc. Natl. Acad. Sci. USA 94, 10862–10867.10.1073/pnas.94.20.10862Suche in Google Scholar PubMed PubMed Central

Yamada, M., Satoh, T., and Mori, M. (2003). Mice lacking the thyrotropin-releasing hormone gene: what do they tell us? Thyroid 13, 1111–1121.10.1089/10507250360731505Suche in Google Scholar PubMed

Supplemental Material:

The online version of this article (DOI: 10.1515/hsz-2015-0192) offers supplementary material, available to authorized users.

Received: 2015-6-10
Accepted: 2015-8-12
Published Online: 2015-8-19
Published in Print: 2016-1-1

©2016 by De Gruyter

Artikel in diesem Heft

  1. Frontmatter
  2. Reviews
  3. Ancestral protein reconstruction: techniques and applications
  4. Mapping the non-standardized biases of ribosome profiling
  5. Minireview
  6. The macromolecular crowding effect – from in vitro into the cell
  7. Research Articles/Short Communications
  8. Protein Structure and Function
  9. IsoQC (QPCTL) knock-out mice suggest differential substrate conversion by glutaminyl cyclase isoenzymes
  10. Molecular Medicine
  11. Correlated overexpression of metadherin and SND1 in glioma cells
  12. Cell Biology and Signaling
  13. Melanoma differentiation-associated gene 5 is involved in the induction of stress granules and autophagy by protonophore CCCP
  14. Suberoylanilide hydroxamic acid (SAHA) promotes the epithelial mesenchymal transition of triple negative breast cancer cells via HDAC8/FOXA1 signals
  15. Melanocytes are more responsive to IFN-γ and produce higher amounts of kynurenine than melanoma cells
  16. Phenobarbital inhibits calpain activity and expression in mouse hepatoma cells
https://doi.org/10.1515/hsz-2015-0192
Schlagwörter für diesen Artikel
5-oxo-L-proline; posttranslational modification; pyroglutamate; TRH

Artikel in diesem Heft

  1. Frontmatter
  2. Reviews
  3. Ancestral protein reconstruction: techniques and applications
  4. Mapping the non-standardized biases of ribosome profiling
  5. Minireview
  6. The macromolecular crowding effect – from in vitro into the cell
  7. Research Articles/Short Communications
  8. Protein Structure and Function
  9. IsoQC (QPCTL) knock-out mice suggest differential substrate conversion by glutaminyl cyclase isoenzymes
  10. Molecular Medicine
  11. Correlated overexpression of metadherin and SND1 in glioma cells
  12. Cell Biology and Signaling
  13. Melanoma differentiation-associated gene 5 is involved in the induction of stress granules and autophagy by protonophore CCCP
  14. Suberoylanilide hydroxamic acid (SAHA) promotes the epithelial mesenchymal transition of triple negative breast cancer cells via HDAC8/FOXA1 signals
  15. Melanocytes are more responsive to IFN-γ and produce higher amounts of kynurenine than melanoma cells
  16. Phenobarbital inhibits calpain activity and expression in mouse hepatoma cells
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