IFN-β induces serine phosphorylation of Stat-1 in Ewing's sarcoma cells and mediates apoptosis via induction of IRF-1 and activation of caspase-7

Four human cell lines derived from Ewing's sarcoma, EW-7, EW-1, COH and ORS, were investigated to establish the effects of human recombinant interferon-alpha 2a and human recombinant interferon-beta on cell proliferation and apoptosis, All four cell lines were much more sensitive to the antiproliferative effects of IFN-beta than of IFN-alpha. Analysis of the early signals triggered by IFN-alpha and IFN-beta demonstrated that the two IFNs were similarly effective in inducing tyrosine phosphorylation of the Jak-1 and Tyk-2 kinases and the transcription factors Stat-1 and Stat-2, Interestingly, an additional rapid phosphorylation of Stat-1 on serine was observed after IFN-beta treatment, with concomitant activation of p38 mitogen-activated protein kinase, In these cells, Stat-1 Ser727 phosphorylation in response to IFN-beta was found to be impaired by p38 MAPkinase inhibitor (SB203580). IFN-beta induced the formation of the Interferon Stimulated Gene Factor 3 complex more efficiently than IFN-alpha, as well as sustained induction of IRF-1, which may account for its greater induction of 2'5'oligo(A)synthetase and greater inhibition of cell proliferation. IFN-beta, but not IFN-alpha, induced apoptosis in wild-type p53 EW-7 and COH cell lines, but not in the mutated p53 EW-1 or ORS cell lines. The apoptosis induced by IFN-beta in EW-7 and COH cell lines appeared to be mediated by IRF-1 and involved the activation of caspase-7, Ectopic expression of IRF-1 induced apoptosis in all four cell lines which correlated with the activation of caspase-7 and,vith the downregulation of the Bcl-2 oncoprotein, as observed for IFN-beta-induced apoptosis in parental EW-7 and COH cell lines.