Keratinocyte growth factor receptor ligands induce transforming growth factor alpha expression and activate the epidermal growth factor receptor signaling pathway in cultured epidermal keratinocytes.

Epidermal growth factor receptor (EGFR) ligands are fundamental regulators of epithelial growth, differentiation, and neoplastic transformation. In addition to being potent mitogens for murine epidermal keratinocytes in vitro, transforming growth factor alpha (TGF alpha) and EGF elicit distinctive changes in keratin expression: Ca(2+)-mediated induction of the differentiation-specific keratins K1 and K10 is blocked, while simple epithelial keratins K8 and K18 are expressed aberrantly (C. Cheng et al., Cell Growth, & Differ., 4: 317-327, 1993). We have evaluated several additional growth factors to determine the specificity of this response for EGFR ligands. TGF alpha, keratinocyte growth factor (KGF), and acidic fibroblast growth factor (aFGF), but not basic fibroblast growth factor (bFGF) or insulin-like growth factor type I, block Ca(2+)-mediated expression of K1 while inducing K8. Since KGF and aFGF (but not bFGF) are ligands for the KGF receptor (KGFR), we explored the possibility that the TGF alpha/EGFR pathway is an intermediary in signaling through the KGFR. TGF alpha mRNA was increased in cells treated with KGF, aFGF, or TGF alpha but not bFGF or insulin-like growth factor type I. Similar changes were detected at the protein level; TGF alpha in conditioned medium (CM) from control, KGF-, TGF alpha-, and aFGF-treated cultures was 54 (+/- 8, SEM), 365 (+/- 50), 146 (+/- 20), and 120 (+/- 50) pg/ml, respectively. KGF and TGF alpha also increased expression of cell-associated TGF alpha measured in keratinocyte lysates. KGF increased TGF alpha secretion and mRNA levels in human as well as mouse keratinocytes. CM from KGF-treated cultures stimulated cell growth when added to cultures of normal keratinocytes. Preincubation with neutralizing antibodies to both TGF alpha and KGF, but not KGF antibody alone, blocked cell growth in cultures treated with KGF CM, suggesting that the predominant keratinocyte mitogen in KGF CM is TGF alpha. In support of this hypothesis, treatment of keratinocytes for 5 min with either KGF CM or purified TGF alpha resulted in EGFR autophosphorylation. Furthermore, after approximately 24 h, KGF as well as TGF alpha induced EGFR down-regulation based on Western blot analysis and 125I-EGF binding. Induction of TGF alpha in KGF-treated keratinocytes, coupled to activation and down-modulation of the EGFR, suggests that TGF alpha may be a proximal effector of KGF action for at least certain aspects of epidermal growth and differentiation.