TGF-beta autocrine loop regulates cell growth and myogenic differentiationin 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 an d myogenic differentiation in the human rhabdomyosarcoma cell, line, RD. We previously reported that the phorbol ester 12-O-tetradecanoylphorbol-13-ac etate (TPA) induces growth arrest and myogenic differentiation in these cel ls, which constitutively express muscle regulatory factors. We show that TP A 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 i s similar to that measured in TPA-treated cells), mimics TPA-induced differ entiation. Taken together, these data demonstrate that cell growth and supp ression 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 my ogenesis 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 prima ry muscle cells a much higher concentration of exogenous TGF-beta is requir ed for the differentiation inhibitory effect and TPA inhibits differentiati on in these cells probably through a TGF-beta independent mechanism. These data thus clarify the mechanism underlying the multiple roles of TGF-beta i n the regulation of both the transformed and differentiated phenotype.-Bouc he, M., Canipari, R., Melchionna, R., Willems, D., Senni, M. I., Molinaro, M. TGF-beta autocrine loop regulates cell growth and myogenic differentiati on in human rhabdomyosarcoma cells.