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Binding to EGF receptor of a laminin-5 EGF-like fragment liberated during MMP-dependent mammary gland involution.

Schenk S, Hintermann E, Bilban M, Koshikawa N, Hojilla C, Khokha R, Quaranta V - J. Cell Biol. (2003)

Bottom Line: Therefore, the elucidation of their identities and functions is of great interest.Here, we show that matrix metalloproteinases (MMPs) generate a domain (DIII) from the ECM macromolecule laminin-5.Binding of a recombinant DIII fragment to epidermal growth factor receptor stimulates downstream signaling (mitogen-activated protein kinase), MMP-2 gene expression, and cell migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. sschenk@scripps

ABSTRACT
Extracellular matrix (ECM) fragments or cryptic sites unmasked by proteinases have been postulated to affect tissue remodeling and cancer progression. Therefore, the elucidation of their identities and functions is of great interest. Here, we show that matrix metalloproteinases (MMPs) generate a domain (DIII) from the ECM macromolecule laminin-5. Binding of a recombinant DIII fragment to epidermal growth factor receptor stimulates downstream signaling (mitogen-activated protein kinase), MMP-2 gene expression, and cell migration. Appearance of this cryptic ECM ligand in remodeling mammary gland coincides with MMP-mediated involution in wild-type mice, but not in tissue inhibitor of metalloproteinase 3 (TIMP-3)-deficient mice, supporting physiological regulation of DIII liberation. These findings indicate that ECM cues may operate via direct stimulation of receptor tyrosine kinases in tissue remodeling, and possibly cancer invasion.

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Related in: MedlinePlus

Induction of EGFR tyrosine phosphorylation by intact Ln-5. Treatment of MDA-MB-231 cells with 1.7 nM EGF for 10 min at 37°C results in significant phosphorylation of 175 kD EGFR (lane 2, left panels) over control (lane 1, no ligand), whereas 2.5 nM purified Ln-5 causes only weak EGFR phosphorylation (lane 3). Stimulation of cells for 90 min (right panels) with Ln-5 (lane 3) results in an EGFR phosphorylation signal well above control (lane 1). In contrast, incubation of cells for 90 min in the presence of EGF (lane 2) diminished the signal toward background level (lane 1).
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fig6: Induction of EGFR tyrosine phosphorylation by intact Ln-5. Treatment of MDA-MB-231 cells with 1.7 nM EGF for 10 min at 37°C results in significant phosphorylation of 175 kD EGFR (lane 2, left panels) over control (lane 1, no ligand), whereas 2.5 nM purified Ln-5 causes only weak EGFR phosphorylation (lane 3). Stimulation of cells for 90 min (right panels) with Ln-5 (lane 3) results in an EGFR phosphorylation signal well above control (lane 1). In contrast, incubation of cells for 90 min in the presence of EGF (lane 2) diminished the signal toward background level (lane 1).

Mentions: To investigate accessibility of DIII for receptor binding within intact Ln-5, MDA-MB-231 cells were incubated with rat Ln-5 under conditions similar to those used for rDIII and EGF. After 10 min, EGFR phosphorylation was stimulated by EGF and to a lesser extent by Ln-5 (Fig. 6) , whereas at 90 min after stimulation, more pronounced receptor phosphorylation was observed with Ln-5 than with EGF. However, Ln-5 did not stimulate ERK1/2 phosphorylation at any time point investigated (5, 20, 30, and 90 min) in either MDA-MB-231 or MCF-7 cells, and this result was independent of whether the cells were in suspension or adherent (unpublished data). Thus, Ln-5–stimulated EGFR phosphorylation differs from that seen with rDIII in at least two respects; slower kinetics and absence of downstream ERK1/2 activation, suggesting that MMP-liberated DIII may have signaling properties distinct from those of intact Ln-5.


Binding to EGF receptor of a laminin-5 EGF-like fragment liberated during MMP-dependent mammary gland involution.

Schenk S, Hintermann E, Bilban M, Koshikawa N, Hojilla C, Khokha R, Quaranta V - J. Cell Biol. (2003)

Induction of EGFR tyrosine phosphorylation by intact Ln-5. Treatment of MDA-MB-231 cells with 1.7 nM EGF for 10 min at 37°C results in significant phosphorylation of 175 kD EGFR (lane 2, left panels) over control (lane 1, no ligand), whereas 2.5 nM purified Ln-5 causes only weak EGFR phosphorylation (lane 3). Stimulation of cells for 90 min (right panels) with Ln-5 (lane 3) results in an EGFR phosphorylation signal well above control (lane 1). In contrast, incubation of cells for 90 min in the presence of EGF (lane 2) diminished the signal toward background level (lane 1).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172889&req=5

fig6: Induction of EGFR tyrosine phosphorylation by intact Ln-5. Treatment of MDA-MB-231 cells with 1.7 nM EGF for 10 min at 37°C results in significant phosphorylation of 175 kD EGFR (lane 2, left panels) over control (lane 1, no ligand), whereas 2.5 nM purified Ln-5 causes only weak EGFR phosphorylation (lane 3). Stimulation of cells for 90 min (right panels) with Ln-5 (lane 3) results in an EGFR phosphorylation signal well above control (lane 1). In contrast, incubation of cells for 90 min in the presence of EGF (lane 2) diminished the signal toward background level (lane 1).
Mentions: To investigate accessibility of DIII for receptor binding within intact Ln-5, MDA-MB-231 cells were incubated with rat Ln-5 under conditions similar to those used for rDIII and EGF. After 10 min, EGFR phosphorylation was stimulated by EGF and to a lesser extent by Ln-5 (Fig. 6) , whereas at 90 min after stimulation, more pronounced receptor phosphorylation was observed with Ln-5 than with EGF. However, Ln-5 did not stimulate ERK1/2 phosphorylation at any time point investigated (5, 20, 30, and 90 min) in either MDA-MB-231 or MCF-7 cells, and this result was independent of whether the cells were in suspension or adherent (unpublished data). Thus, Ln-5–stimulated EGFR phosphorylation differs from that seen with rDIII in at least two respects; slower kinetics and absence of downstream ERK1/2 activation, suggesting that MMP-liberated DIII may have signaling properties distinct from those of intact Ln-5.

Bottom Line: Therefore, the elucidation of their identities and functions is of great interest.Here, we show that matrix metalloproteinases (MMPs) generate a domain (DIII) from the ECM macromolecule laminin-5.Binding of a recombinant DIII fragment to epidermal growth factor receptor stimulates downstream signaling (mitogen-activated protein kinase), MMP-2 gene expression, and cell migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. sschenk@scripps

ABSTRACT
Extracellular matrix (ECM) fragments or cryptic sites unmasked by proteinases have been postulated to affect tissue remodeling and cancer progression. Therefore, the elucidation of their identities and functions is of great interest. Here, we show that matrix metalloproteinases (MMPs) generate a domain (DIII) from the ECM macromolecule laminin-5. Binding of a recombinant DIII fragment to epidermal growth factor receptor stimulates downstream signaling (mitogen-activated protein kinase), MMP-2 gene expression, and cell migration. Appearance of this cryptic ECM ligand in remodeling mammary gland coincides with MMP-mediated involution in wild-type mice, but not in tissue inhibitor of metalloproteinase 3 (TIMP-3)-deficient mice, supporting physiological regulation of DIII liberation. These findings indicate that ECM cues may operate via direct stimulation of receptor tyrosine kinases in tissue remodeling, and possibly cancer invasion.

Show MeSH
Related in: MedlinePlus