A role for cathepsin E in the processing of mast-cell carboxypeptidase A

J Cell Sci. 2005 May 1;118(Pt 9):2035-42. doi: 10.1242/jcs.02333.

Abstract

Mast-cell carboxypeptidase A is stored in the secretory granule and is released, together with a range of other inflammatory mediators, upon mast-cell degranulation. Carboxypeptidase A, like all mast-cell proteases, is stored in the granule as an active enzyme (i.e. with its propeptide removed). Although the processing mechanisms for the other classes of mast-cell proteases (in particular the chymases) have been clarified to some extent, the processing of procarboxypeptidase A is poorly characterized. Here, we show that mast cells from mice lacking the aspartic protease cathepsin E display an accumulation of procarboxypeptidase A, indicating a defect in carboxypeptidase-A processing. By contrast, mast cells lacking cathepsins B, L or D have normal carboxypeptidase-A processing. Furthermore, recombinant cathepsin E was found to process recombinant procarboxypeptidase A in vitro, under conditions resembling those found in mast-cell granules. Immunohistochemical analysis revealed staining for cathepsin E in mast cells from normal mice but not in mast cells from mice lacking heparin, indicating that cathepsin E is bound to heparin proteoglycan within mast-cell granules. In accordance with this notion, affinity chromatography showed that recombinant cathepsin E bound strongly to heparin under acidic conditions (the conditions prevailing in mast-cell granules) but not at neutral pH. Moreover, mast-cell degranulation resulted in the release of cathepsin E. Taken together, our results indicate that cathepsin E is located in mast-cell secretory granules in complex with heparin proteoglycans, and that it has a role in the processing of procarboxypeptidase A into active protease.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Carboxypeptidases A / chemistry
  • Carboxypeptidases A / metabolism*
  • Cathepsin B / physiology
  • Cathepsin D / physiology
  • Cathepsin E / chemistry
  • Cathepsin E / metabolism
  • Cathepsin E / physiology*
  • Cathepsin L
  • Cathepsins / physiology
  • Cell Line
  • Cells, Cultured
  • Cloning, Molecular
  • Cysteine Endopeptidases / physiology
  • Enzyme Activation
  • Female
  • Heparin / chemistry
  • Humans
  • Hydrogen-Ion Concentration
  • Immunohistochemistry
  • Inflammation
  • Mast Cells / enzymology*
  • Mice
  • Mice, Inbred C57BL
  • Models, Biological
  • Models, Molecular
  • Peptide Hydrolases / metabolism
  • Proteoglycans / chemistry
  • Recombinant Proteins / chemistry
  • Secretory Vesicles / metabolism
  • Time Factors

Substances

  • Proteoglycans
  • Recombinant Proteins
  • Heparin
  • Cathepsins
  • Peptide Hydrolases
  • CPA3 protein, human
  • Carboxypeptidases A
  • Cpa3 protein, mouse
  • Cysteine Endopeptidases
  • Cathepsin B
  • CTSL protein, human
  • Cathepsin L
  • Ctsl protein, mouse
  • Cathepsin E
  • Cathepsin D