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TIMP-2 is released as an intact molecule following binding to MT1-MMP on the cell surface

Stanley Zucker1, Michelle Hymowitz, Cathleen Conner

  • 1Department of Research, Veterans Affairs Medical Center, Northport, NY 11768, USA. s_zucker@yahoo.com

Experimental Cell Research
|January 20, 2004
PubMed

Insights

Tissue inhibitor of metalloproteinase-2 (TIMP-2) primarily returns to circulation intact after binding to membrane type-1 MMP (MT1-MMP). This study re-examines TIMP-2

Area of Science:

  • Biochemistry
  • Cell Biology
  • Enzymology

Background:

  • Matrix metalloproteinase-2 (MMP-2) activation requires binding of tissue inhibitor of metalloproteinase-2 (TIMP-2) to pro-MMP-2 and membrane type-1 MMP (MT1-MMP) on the cell surface.
  • Previous studies suggested TIMP-2 undergoes extensive lysosomal degradation after cell surface receptor binding and endocytosis.

Purpose of the Study:

  • To re-examine the fate of TIMP-2 after binding to cell surface MT1-MMP.
  • To investigate the release, endocytosis, and degradation of TIMP-2 under various conditions.

Main Methods:

  • Utilized radiolabeled 125I-TIMP-2 binding assays on transfected HT1080 cells expressing MT1-MMP.
  • Employed dominant-negative dynamin cDNA transfection, pharmacological agents (concanavalin A, bafilomycin A1), and a furin inhibitor to modulate cellular processes.
  • Confirmed MT1-MMP presence and maturation using biotinylation and Western blotting.

Main Results:

  • Over 85% of bound 125I-TIMP-2 was released as intact molecules, with less than 15% degraded.
  • Inhibition of endocytosis (dynamin mutant) delayed TIMP-2 release.
  • Receptor clustering and lysosomal inhibition enhanced TIMP-2 binding.
  • Furin inhibition blocked MT1-MMP activation, preventing TIMP-2 binding and endocytosis.

Conclusions:

  • TIMP-2 is primarily released from cells as an intact, functional molecule after binding to cell surface MT1-MMP.
  • Endocytosis plays a role in TIMP-2 processing, but degradation is limited.
  • MT1-MMP activation is crucial for TIMP-2 interaction and subsequent cellular fate.

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