PAI-1 mediates the TGF-Β1EGF-induced scatter response in transformed human keratinocytes

Abstract

Cooperative interactions between growth factor signaling pathways are important elements in carcinoma progression. A model system combining transforming growth factor-Β1 (TGF-Β1) and EGF was developed to investigate mechanisms underlying induced epithelial-to-mesenchymal transition (EMT) in ras-transformed human (HaCaT II-4) keratinocytes. Dual stimulation with TGF-Β1EGF resulted in keratinocyte plasticity and pronounced colony dispersal. The most highly expressed transcript, identified by mRNA profiling, encoded plasminogen activator inhibitor-1 (PAI-1; SERPINE1). PAI-1 negatively regulates plasmin-dependent matrix degradation, preserving a stromal scaffold permissive for keratinocyte motility. Mitogen-activated extracellular kinase (MEK)/extracellular signal-regulated kinase (ERK) and p38 signaling were required for maximal PAI-1 upregulation and TGF-Β+1EGF-stimulated cell locomotion, as pharmacologic disruption of MEK/p38 activity ablated both responses. Moreover, PAI-1 knockdown alone effectively inhibited TGF-Β1EGF-dependent cell scattering, indicating a functional role for this SERPIN in the dual-growth factor model of induced motility. Moreover, EGFR signaling blockade or EGFR knockdown attenuated TGF-Β1-induced PAI-1 expression, implicating EGFR transactivation in TGF-Β1-stimulated PAI-1 expression, and reduced colony dispersal in TGF-Β1EGF-treated cultures. Identification of such cooperative signaling networks and their effect on specific invasion-promoting target genes, such as PAI-1, may lead to the development of pathway-specific therapeutics that affect late-stage events in human tumor progression. © 2010 The Society for Investigative Dermatology.