TGF-beta autocrine loop regulates cell growth and myogenic differentiation in human rhabdomyosarcoma cells.
Transforming growth factor beta (TGF) is a well-known inhibitor of myogenic differentiation as well as an autocrine product of rhabdomyosarcoma cells. We studied the role of the TGF-beta autocrine loop in regulating growth and myogenic differentiation in the human rhabdomyosarcoma cell line, RD. We previously reported that the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) induces growth arrest and myogenic differentiation in these cells, which constitutively express muscle regulatory factors. We show that TPA inhibits the activation of secreted latent TGF-beta, thus decreasing the concentration of active TGF-beta to which the cells are exposed. This event is mediated by the TPA-induced alteration of the uPA/PAI serine-protease system. Complete removal of TGF-beta, mediated by the ectopic expression of a soluble type II TGF-beta receptor dominant negative cDNA, induces growth arrest, but does not trigger differentiation. In contrast, a reduction in the TGF-beta concentration, to a range of 0.14-0.20 x 10(-2) ng/ml (which is similar to that measured in TPA-treated cells), mimics TPA-induced differentiation. Taken together, these data demonstrate that cell growth and suppression of differentiation in rhabdomyosarcoma cells require overproduction of active TGF-beta; furthermore, they show that a 'critical' concentration of TGF-beta is necessary for myogenic differentiation to occur, whereas myogenesis is abolished below and above this concentration. By impairing the TGF-beta autocrine loop, TPA stabilizes the factor concentration within the range compatible for differentiation to occur. In contrast, in human primary muscle cells a much higher concentration of exogenous TGF-beta is required for the differentiation inhibitory effect and TPA inhibits differentiation in these cells probably through a TGF-beta independent mechanism. These data thus clarify the mechanism underlying the multiple roles of TGF-beta in the regulation of both the transformed and differentiated phenotype.
Bouché
M
M
Department of Histology and Medical Embryology, University of Rome 'La Sapienza', 00161, Rome, Italy. bouche@uniroma1.it
Canipari
R
R
Melchionna
R
R
Willems
D
D
Senni
M I
MI
Molinaro
M
M
D000818:Animals; D007611:Aprotinin / Q000494:pharmacology; D019898:Autocrine Communication / Q000187:drug effects*; D002454:Cell Differentiation / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D006801:Humans; D018482:Muscle, Skeletal / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D009154:Mutation / Q000235:genetics; D018995:Myosin Heavy Chains / Q000378:metabolism; D010436:Pepstatins / Q000494:pharmacology; D010958:Plasminogen / Q000378:metabolism; D015849:Plasminogen Inactivators / Q000378:metabolism; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism; D012208:Rhabdomyosarcoma / Q000378:metabolism / Q000473:pathology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors / Q000378:metabolism
D010436:Pepstatins; D015849:Plasminogen Inactivators; D011498:Protein Precursors; D012333:RNA, Messenger; D018125:Receptors, Transforming Growth Factor beta; D016212:Transforming Growth Factor beta; C014195:Streptomyces pepsin inhibitor; D010958:Plasminogen; D007611:Aprotinin; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II; D014568:Urokinase-Type Plasminogen Activator; D018995:Myosin Heavy Chains; D013755:Tetradecanoylphorbol Acetate; C031375:pepstatin