Membrane-associated HB-EGF modulates HGF-induced cellular responses in MDCK cells

Abstract

In MDCK cells, hepatocyte growth factor/scatter factor (HGF/SF) induces epithelial cell dissociation, scattering, migration, growth and formation of branched tubular structures. By contrast, these cells neither scatter nor form tubular structures in response to the epidermal growth factor (EGF) family of growth factors. Heparin-binding EGF-like growth factor (HB-EGF) is a member of the EGF family of growth factors and is synthesized as a membrane-associated precursor molecule (proHB-EGF). ProHB-EGF is proteolytically cleaved to release a soluble ligand (sHB-EGF) that activates the EGF receptor. Although recent studies suggest possible physiological functions, the role of proHB-EGF remains largely undefined. Using MDCK cells stably expressing proHB-EGF, a noncleavable deletion mutant of proHB-EGF or soluble HB-EGF, we show that epithelial cell functions differ depending on the form of HB-EGF being expressed. Expression of noncleavable membrane-anchored HB-EGF promoted cell-matrix and cell-cell interactions and decreased cell migration, HGF/SF-induced cell scattering and formation of tubular structures. By contrast, expression of soluble HB-EGF induced increased cell migration, decreased cell-matrix and cell-cell interactions and promoted the development of long unbranched tubular structures in response to HGF/SF. These findings suggest that HB-EGF can not only modulate HGF/SF-induced cellular responses in MDCK cells but also that membrane-bound HB-EGF and soluble HB-EGF give rise to distinctly different effects on cell-cell and cell-extracellular matrix interactions.

Soluble HB-EGF induces epithelial-to-mesenchymal transition in inner medullary collecting duct cells by upregulating Snail-2

“…1B). These data are consistent with chronic activation of the EGFR; ligand-induced EGFR activation induces internalization/degradation of the receptor (6,38). IMCD sHB cells demonstrated a marked change in morphology compared with vector-transfected IMCD cells (IMCD vec ): they did not grow in epithelial sheets, and they exhibited significant cytoplasmic projections (Fig.…”

“…To determine whether increased expression of Snail-2 in response to sHB-EGF was a cell line-specific phenomenon, we assessed its expression in MDCK II cells that were stably transfected to overexpress sHB-EGF (38). Similar to our results with IMCD cells, MDCK sHB cells had increased expression of Snail-2 compared with MDCK vec (Fig.…”

“…5C). As previously reported, MDCK sHB cells exhibit increased cell migration and decreased cell-matrix and cell-cell interactions; in addition, these cells develop tubular structures in response to hepatocyte growth factor (38). Interestingly, overexpression of sHB-EGF does not lead to a detectable decrease in E-cadherin protein expression in MDCK II cells, however.…”

“…To develop IMCD cells that stably express soluble HB-EGF, we followed the same procedure that we previously described for the creation of sHB-EGF-overexpressing MDCK II cells (38). Briefly, IMCD cells were transfected with 1 g of pcDNA3-neo containing rat sHB-EGF cDNA (38) using Lipofectamine reagent (Invitrogen). After five passages in DMEM/F12 containing 1 mg/ml G418, stable transfectants were selected with cloning cylinders and subcloned by serial dilution.…”

Regulation of Cell Motility by Tyrosine Phosphorylated Villin

“…Similarly, EGF has been reported to enhance epithelial cell migration by mechanisms varied from HGF (Singh et al, 2004) as well as comparable with HGF (Muller et al, 2002). We measured the effects of both HGF and EGF on villin-induced cell migration.…”