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Identification of ADAM10 as a major TNF sheddase in ADAM17-deficient fibroblasts

Renata Mezyk-Kopeć1, Monika Bzowska, Krystyna Stalińska

  • 1Department of Cell Biochemistry, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, ul. Gronostajowa 7, 30-387 Kraków, Poland.

Cytokine
|April 7, 2009
PubMed

Insights

ADAM17-deficient cells shed soluble tumor necrosis factor (TNF) via a metalloprotease. This activity, inhibited by specific agents and stimulated by ionomycin, matches ADAM10, suggesting ADAM10 acts as the TNF sheddase in ADAM17-deficient cells.

Area of Science:

  • Biochemistry
  • Cell Biology
  • Immunology

Background:

  • Tumor necrosis factor (TNF) is a crucial cytokine involved in inflammation and immunity.
  • TNF shedding from the cell membrane is a critical step in its activation and signaling.
  • The metalloprotease ADAM17 (a disintegrin and metalloprotease) is known to be a primary TNF sheddase.

Purpose of the Study:

  • To investigate the enzyme responsible for TNF shedding in murine fibroblasts deficient in ADAM17.
  • To characterize the enzymatic activity and identify the specific metalloprotease involved in proTNF processing.

Main Methods:

  • Utilized ADAM17-deficient murine fibroblasts stably transfected with proTNF cDNA.
  • Assessed TNF shedding activity and its inhibition by specific metalloprotease inhibitors (1,10-phenantroline, GM6001) and stimulators (ionomycin).
  • Investigated the role of ADAM10 by employing shRNA-mediated silencing and analyzing ADAM10 expression levels.

Main Results:

  • ADAM17-deficient cells released biologically active soluble TNF, indicating the presence of an alternative sheddase.
  • The enzyme responsible hydrolyzed proTNF at the Ala(76):Val(77) bond, with activity profiles matching ADAM10.
  • Moderate silencing of ADAM10 significantly inhibited TNF shedding, and no correlation was found between ADAM10 expression and active ADAM17 presence.

Conclusions:

  • ADAM10 can function as a TNF sheddase in cells lacking ADAM17 activity.
  • This finding reveals a potential compensatory mechanism for TNF processing in specific cellular contexts.
  • Highlights the distinct roles and potential interplay between ADAM10 and ADAM17 in regulating TNF bioavailability.

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