Transforming growth factor-beta-induced mobilization of actin cytoskeleton required signaling by small GTPases Cdc42 and RhoA

• Transforming growth factor-beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. We studied TGF-beta-induced rearrangements of the actin filament system and found that TGF-beta 1 treatment of PC-3U human prostate carcinoma cells resulted in a rapid formation of lamellipodia. Interestingly, this response was shown to be independent of the Smad signaling pathway; instead, it required the activity of the Rho GTPases Cdc42 and RhoA, because ectopic expression of dominant negative mutant Cdc42 and RhoA abrogated the response. Long-term stimulation with TGF-beta 1 resulted in an assembly of stress fibers; this response required both signaling via Cdc42 and RhoA, and Smad proteins. A known downstream effector of Cdc42 is p38(MAPK); treatment of the cells with the p38(MAPK) inhibitor 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(pyridyl)1H-imidazole (SB203580), as well as ectopic expression of a kinase-inactive p38(MAPK), abrogated the TGF-beta-induced actin reorganization. Moreover, treatment of cells with the inhibitors of the RhoA target-protein Rho-associated coiled-coil kinase (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide (Y-27632) and 1-5(-isoquinolinesulfonyl)homopiperazine (HA-1077), as well as ectopic expression of kinase-inactive Rho coiled-coil kinase-1, abrogated the TGF-beta 1-induced formation of stress fibers. Collectively, these data indicate that TGF-beta-induced membrane ruffles occur via Rho GTPase-dependent pathways, whereas long-term effects require cooperation between Smad and Rho GTPase signaling pathways.

Nyckelord

Actins/*metabolism

Amides/pharmacology

Animals

Cell Surface Extensions/metabolism

Cytoskeleton/drug effects/*metabolism

DNA-Binding Proteins/genetics/metabolism

Enzyme Inhibitors/pharmacology

Humans

Imidazoles/pharmacology

Intracellular Signaling Peptides and Proteins

Mitogen-Activated Protein Kinases/antagonists & inhibitors/genetics/metabolism

Protein-Serine-Threonine Kinases/antagonists & inhibitors/genetics/metabolism

Pyridines/pharmacology

Rats

Recombinant Fusion Proteins/metabolism

Signal Transduction/*physiology

Smad4 Protein

Stress Fibers/metabolism

Trans-Activators/genetics/metabolism

Transforming Growth Factor beta/*metabolism

Tumor Cells; Cultured

cdc42 GTP-Binding Protein/*metabolism

p38 Mitogen-Activated Protein Kinases

rac1 GTP-Binding Protein/metabolism

rhoA GTP-Binding Protein/*metabolism

MEDICINE

MEDICIN

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