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Expression and function of matrix metalloproteinase (MMP)-28.

Rodgers UR, Kevorkian L, Surridge AK, Waters JG, Swingler TE, Culley K, Illman S, Lohi J, Parker AE, Clark IM - Matrix Biol. (2009)

Bottom Line: Adhesion to type II collagen and fibronectin was increased, and migration across the former was decreased.These data demonstrate that expression of MMP28 alters cell phenotype towards a more adhesive, less migratory behaviour.Further, MMP-28 activity may reside predominantly in the extracellular matrix, and we are currently searching for substrates in this compartment.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Research Centre, School of Biological Sciences, University of East Anglia, Norwich, UK.

ABSTRACT
Matrix metalloproteinase-28 (MMP-28, epilysin) is highly expressed in the skin by keratinocytes, the developing and regenerating nervous system and a number of other normal human tissues. In epithelial cells, over-expression of MMP-28 mediates irreversible epithelial to mesenchymal transition concomitant with loss of E-cadherin from the cell surface and an increase in active transforming growth factor beta. We recently reported the expression of MMP-28 in both cartilage and synovium where expression is increased in patients with osteoarthritis. In human chondrosarcoma cells MMP-28 was activated by proprotein convertases and the active form of the enzyme preferentially associated with the extracellular matrix in a C-terminal independent manner. over-expression of MMP-28 in chondrosarcoma cells led to altered cell morphology with increased organisation of actin. Adhesion to type II collagen and fibronectin was increased, and migration across the former was decreased. MMP-28 was localised to the cell surface, at least transiently, in a C-terminal dependent manner. Heparin prevented both extracellular matrix association and cell surface binding of MMP-28 suggesting that both are via heparan sulphate proteoglycans. Over-expression of activatable MMP-28, but not catalytically inactive EA mutant increased the expression and activity of MMP-2, and all forms of MMP-28 tested increased expression of MMP19 and TIMP3 mRNA. These data demonstrate that expression of MMP28 alters cell phenotype towards a more adhesive, less migratory behaviour. Further, MMP-28 activity may reside predominantly in the extracellular matrix, and we are currently searching for substrates in this compartment.

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Heparin competes for cell surface-associated MMP-28. SW1353 cells stably transfected with the wild-type MMP28 expression construct were probed, non-permeabilised, for protein expression with an anti-FLAG primary antibody and an Alexa 488-conjugated secondary antibody. Cells were either untreated (A, D) or treated with 0.1 mg/ml heparin (B, E). Wild-type cells treated with an IgG isotype control primary antibody were also included as a control (C). Cells were counterstained with DAPI and viewed under 20× magnification.
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fig5: Heparin competes for cell surface-associated MMP-28. SW1353 cells stably transfected with the wild-type MMP28 expression construct were probed, non-permeabilised, for protein expression with an anti-FLAG primary antibody and an Alexa 488-conjugated secondary antibody. Cells were either untreated (A, D) or treated with 0.1 mg/ml heparin (B, E). Wild-type cells treated with an IgG isotype control primary antibody were also included as a control (C). Cells were counterstained with DAPI and viewed under 20× magnification.

Mentions: The addition of heparin to the cultured wild-type transfects (as above) led to a reduction in cell surface staining for MMP-28 in non-permeabilised cells (Fig. 5), demonstrating competition for binding sites in a similar manner to that seen for ECM binding.


Expression and function of matrix metalloproteinase (MMP)-28.

Rodgers UR, Kevorkian L, Surridge AK, Waters JG, Swingler TE, Culley K, Illman S, Lohi J, Parker AE, Clark IM - Matrix Biol. (2009)

Heparin competes for cell surface-associated MMP-28. SW1353 cells stably transfected with the wild-type MMP28 expression construct were probed, non-permeabilised, for protein expression with an anti-FLAG primary antibody and an Alexa 488-conjugated secondary antibody. Cells were either untreated (A, D) or treated with 0.1 mg/ml heparin (B, E). Wild-type cells treated with an IgG isotype control primary antibody were also included as a control (C). Cells were counterstained with DAPI and viewed under 20× magnification.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2724077&req=5

fig5: Heparin competes for cell surface-associated MMP-28. SW1353 cells stably transfected with the wild-type MMP28 expression construct were probed, non-permeabilised, for protein expression with an anti-FLAG primary antibody and an Alexa 488-conjugated secondary antibody. Cells were either untreated (A, D) or treated with 0.1 mg/ml heparin (B, E). Wild-type cells treated with an IgG isotype control primary antibody were also included as a control (C). Cells were counterstained with DAPI and viewed under 20× magnification.
Mentions: The addition of heparin to the cultured wild-type transfects (as above) led to a reduction in cell surface staining for MMP-28 in non-permeabilised cells (Fig. 5), demonstrating competition for binding sites in a similar manner to that seen for ECM binding.

Bottom Line: Adhesion to type II collagen and fibronectin was increased, and migration across the former was decreased.These data demonstrate that expression of MMP28 alters cell phenotype towards a more adhesive, less migratory behaviour.Further, MMP-28 activity may reside predominantly in the extracellular matrix, and we are currently searching for substrates in this compartment.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Research Centre, School of Biological Sciences, University of East Anglia, Norwich, UK.

ABSTRACT
Matrix metalloproteinase-28 (MMP-28, epilysin) is highly expressed in the skin by keratinocytes, the developing and regenerating nervous system and a number of other normal human tissues. In epithelial cells, over-expression of MMP-28 mediates irreversible epithelial to mesenchymal transition concomitant with loss of E-cadherin from the cell surface and an increase in active transforming growth factor beta. We recently reported the expression of MMP-28 in both cartilage and synovium where expression is increased in patients with osteoarthritis. In human chondrosarcoma cells MMP-28 was activated by proprotein convertases and the active form of the enzyme preferentially associated with the extracellular matrix in a C-terminal independent manner. over-expression of MMP-28 in chondrosarcoma cells led to altered cell morphology with increased organisation of actin. Adhesion to type II collagen and fibronectin was increased, and migration across the former was decreased. MMP-28 was localised to the cell surface, at least transiently, in a C-terminal dependent manner. Heparin prevented both extracellular matrix association and cell surface binding of MMP-28 suggesting that both are via heparan sulphate proteoglycans. Over-expression of activatable MMP-28, but not catalytically inactive EA mutant increased the expression and activity of MMP-2, and all forms of MMP-28 tested increased expression of MMP19 and TIMP3 mRNA. These data demonstrate that expression of MMP28 alters cell phenotype towards a more adhesive, less migratory behaviour. Further, MMP-28 activity may reside predominantly in the extracellular matrix, and we are currently searching for substrates in this compartment.

Show MeSH
Related in: MedlinePlus