Obligatory role for phospholipase C-γ₁ in villin-induced epithelial cell migration

Abstract

While there is circumstantial evidence to suggest a requirement for phospholipase C-γ1 (PLC-γ1) in actin reorganization and cell migration, few studies have examined the direct mechanisms that link regulators of the actin cytoskeleton with this crucial signaling molecule. This study was aimed to examine the role that villin, an epithelial cell-specific actin-binding protein, and its ligand PLC-γ1 play in migration in intestinal and renal epithelial cell lines that endogenously or ectopically express human villin. Basal as well as epidermal growth factor (EGF)-stimulated cell migration was accompanied by tyrosine phosphorylation of villin and its association with PLC-γ1. Inhibition of villin phosphorylation prevented villin-PLC-γ1 complex formation as well as villin-induced cell migration. The absolute requirement for PLC-γ1 in villin-induced cell migration was demonstrated by measuring cell motility in PLC-γ1−/− cells and by downregulation of endogenous PLC-γ1. EGF-stimulated direct interaction of villin with the Src homology domain 2 domain of PLC-γ1 at the plasma membrane was demonstrated in living cells by using fluorescence resonance energy transfer. These results demonstrate that villin provides an important link between the activation of phosphoinositide signal transduction pathway and epithelial cell migration.

A Novel Role for Villin in Intestinal Epithelial Cell Survival and Homeostasis

“…We have previously demonstrated the induction of villin following doxycycline withdrawal and shown the absence of villin in the MDCK and HeLa Tet-Off cells in the presence of doxycycline (16,17). In addition, IEC-6 cells were infected with recombinant adenovirus expressing full-length human villin protein as described previously (19). To induce cell death, cells were treated with camptothecin (20 M), staurosporine (1 M), a combination of TNF-␣ (10 ng/ml) and cycloheximide (10 ng/ml), or serum-starved for 72 h.…”

Regulation of directional cell migration by membrane-induced actin bundling

“…For all studies with transfected cells, multiple clones of each cell line with similar protein expression levels or a mixed clone were used to avoid clonal variations. MDCK Tet-Off and Caco-2 BBe1 cells were cultured as described previously (Tomar et al, 2006;Wang et al, 2007).…”

The role of actin bundling proteins in the assembly of filopodia in epithelial cells

“…99,106,[136][137][138][139][140][141][142] Villin is unique among the actin-bundling proteins and in fact among all actinbinding proteins in that it can cap, sever, nucleate and bundle actin filaments. These unique properties of villin may contribute to the diverse functions performed by villin in epithelial cells.…”