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Metalloproteinase-mediated release of human Fas ligand
N Kayagaki1, A Kawasaki, T Ebata
1Department of Immunology, Juntendo University School of Medicine, Tokyo, Japan.
Abstract:
Fas ligand (FasL) is a type II integral membrane protein homologous with tumor necrosis factor (TNF). Recent studies indicate that TNF is processed to yield the soluble cytokine by metalloproteinases at the cell surface of activated macrophages and T cells. In the present study, we investigated whether FasL is also released by metalloproteinases. Treatment with hydroxamic acid inhibitors of matrix metalloproteinases specifically led to accumulation of membrane-type FasL (p40) on the surface of human FasL cDNA transfectants and activated human T cells, as estimated by surface immunofluorescence and immunoprecipitation with newly established anti-human FasL monoclonal antibodies. This surface accumulation of mFasL was associated with the decrease of soluble FasL (p27) in the supernatant as estimated by quantitative ELISA and immunoprecipitation with anti-human FasL monoclonal antibodies. These results indicate that human FasL is efficiently released from the cell surface by metalloproteinases like TNF.
Insights
Metalloproteinases release soluble Fas ligand (FasL) from cell surfaces, similar to tumor necrosis factor (TNF). Inhibiting these enzymes increases membrane-bound FasL and decreases soluble FasL, indicating metalloproteinase-mediated shedding.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Fas ligand (FasL) is a type II integral membrane protein homologous to tumor necrosis factor (TNF).
- Tumor necrosis factor (TNF) is known to be processed and released as a soluble cytokine by metalloproteinases.
- The mechanism of FasL release from cell surfaces was not fully understood.
Purpose of the Study:
- To investigate whether FasL is released from the cell surface by metalloproteinases.
- To determine the role of metalloproteinases in the processing and shedding of FasL.
Main Methods:
- Treatment of human FasL cDNA transfectants and activated human T cells with hydroxamic acid inhibitors of matrix metalloproteinases.
- Surface immunofluorescence and immunoprecipitation using anti-human FasL monoclonal antibodies to estimate membrane-type FasL (p40).
- Quantitative ELISA and immunoprecipitation to measure soluble FasL (p27) in the supernatant.
Main Results:
- Inhibitors of matrix metalloproteinases caused an accumulation of membrane-type FasL (p40) on the cell surface.
- This accumulation of membrane-bound FasL was accompanied by a decrease in soluble FasL (p27) in the supernatant.
- These findings suggest a direct role for metalloproteinases in FasL release.
Conclusions:
- Human FasL is efficiently released from the cell surface by metalloproteinases.
- This metalloproteinase-mediated shedding of FasL is analogous to the processing of TNF.
- The study elucidates a key mechanism in the regulation of FasL bioavailability and function.