Hepatocyte growth factor induces MAPK-dependent formin IV translocation inrenal epithelial cells

Renal epithelial tubule formation in cultured cells occurs after the additi on of tubulogenic growth factors such as the hepatocyte growth factor (HGF) . HGF activates the tyrosine kinase receptor c-met, initiating a series of complex events that regulate cell morphology, cell-cell interactions, and c ell-matrix interactions and eventually result in the formation of branching tubular structures. The discovery that disruption of the formin gene locus in mice causes agenesis of the kidneys secondary to failure of ureteric bu d outgrowth and branching tubule formation suggested that this family of pr oteins may be critical to the development of renal epithelial tubules. In t his study, we investigated whether formin is involved in the HGF/c-met sign aling pathway of in vitro tubulogenesis in renal epithelial cells. mIMCD-3 cells were analyzed by reverse transcription-PCR and found to express formi n IV mRNA. With the use of an antibody that recognizes the carboxy terminus of all known formin isoforms, it was observed a formin isoform of approxim ately 165 kD markedly increased in the detergent soluble cell lysate after 10 min of stimulation with HGF. An antibody that is specific for formin IV was then generated and confirmed that the formin isoform regulated by HGF w as formin IV. Cell fractionation and confocal localization of formin IV rev ealed that formin IV is primarily found in a submembranous band that co-loc alizes with the actin cytoskeleton and in a perinuclear location in quiesce nt epithelial cells but undergoes a rapid relocalization after HGF stimulat ion with translocation into the cell cytosol and into the nucleus. Formin T V was found to be a phosphorylation substrate for activated extracellular s ignal-regulated kinase in vitro, and pretreatment of cells with the mitogen -activated protein kinase inhibitor U0126 prevented the translocation of fo rmin IV and inhibited HGF-dependent phosphorylation of formin IV in intact cells. In conclusion, activation of the c-met receptor results in cellular relocalization of formin TV in a mitogen-activated protein kinase-dependent manner.