Regulation of cell surface APO-1/Fas (CD95) ligand expression by metalloproteases

Eur J Immunol. 1995 Aug;25(8):2303-7. doi: 10.1002/eji.1830250828.

Abstract

APO-1/Fas (CD95) ligand (APO-1L) induces apoptosis in sensitive target cells. Activation-induced T cell death and Ca2(+)-independent cytotoxicity in perforin knockout mice are mediated by APO-1L. To define whether APO-1L is expressed on the surface of activated T cells and to investigate the mechanisms leading to the release of a soluble form, we developed rabbit anti-APO-1L antibodies (Ab). The purified rabbit Ab detected the mature forms of the human and mouse APO-1L of approximately 42 and 40 kDa. In addition, the Ab recognized the non-glycosylated form of APO-1L of approximately 32-33 kDa. In activated human T cells, the soluble form of APO-1L was detectable with a molecular mass of 26 kDa. Immunofluorescence of three human T lymphoblastoid cell lines showed that activation of these cells by phorbol 12-myristate 13-acetate/ionomycin induced a significant increase in cell surface APO-1L only in the presence of metalloprotease inhibitors. Zn2+, but not Ca2+, prevented the increase in surface APO-1L observed in the presence of 1,10-phenanthroline. Blocking of other classes of proteases (serine- and acid-proteases, chymotrypsin) had no effect. Increased expression of surface APO-1L by metalloprotease inhibitors was not dependent on T cell activation, as the metalloprotease inhibitors also modulated the low level of constitutive APO-1L expression. These results suggest that cell surface expression of human APO-1L is regulated by Zn2(+)-dependent metalloproteases. Cleavage of surface APO-1L may act as a regulatory mechanism to prevent accumulation of the membrane-bound form and may cause systemic effects of the APO-1L.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, Surface / biosynthesis*
  • Antigens, Surface / genetics*
  • Antigens, Surface / immunology
  • Base Sequence
  • Blotting, Western
  • Cell Line
  • Gene Expression Regulation
  • Humans
  • Immune Sera / immunology
  • Lymphocyte Activation / immunology
  • Metalloendopeptidases / antagonists & inhibitors
  • Metalloendopeptidases / physiology*
  • Molecular Sequence Data
  • Rabbits
  • T-Lymphocytes / metabolism*
  • Up-Regulation
  • fas Receptor

Substances

  • Antigens, Surface
  • Immune Sera
  • fas Receptor
  • Metalloendopeptidases