TUMOR-NECROSIS-FACTOR-ALPHA UP-REGULATES GI-ALPHA AND G-BETA PROTEINS AND ADENYLYL CYCLASE RESPONSIVENESS IN RAT CARDIOMYOCYTES

Treatment of cultured rat cardiomyocytes in serum-free medium for 48 h with recombinant human tumor necrosis factor-alpha (TNF-alpha) led to a concentration-dependent increase in the level of membrane-inhibitory guanine nucleotide-binding protein (G(i)) alpha-subunits and in pertussis toxin-catalyzed [P-32]ADP ribosylation of 40 kDa proteins. Both G(i-alpha) protein subtypes present in rat cardiac myocyte membranes, G(i-alpha-40) and G(i-alpha-41), were up-regulated by the cytokine, with the maximal increase occurring at 10 U/ml TNF-alpha. In contrast to noradrenaline exposure, which causes a similar, but apparently exclusive, increase in alpha-i-subunits, treatment with TNF-alpha in addition increased the level of membrane G protein beta-36-subunits. Furthermore, while noradrenaline exposure led to a decrease in receptor-dependent and -independent adenylyl cyclase activity, treatment of cardiomyocytes with TNF-alpha caused a concentration-dependent increase in cyclase responsiveness to either forskolin, guanosine 5'-O-(3-thiotriphosphate) or isoproterenol, even though beta-adrenoceptor density was decreased by TNF-alpha. The increase in adenylyl cyclase activity induced by TNF-alpha was completely suppressed when the cells were cocultured with noradrenaline, a condition leading to an additive increase in G(i-alpha) level. The data indicate that the cytokine TNF-alpha can potently modulate G protein-mediated signal transduction in rat cardiac myocytes. Although TNF-alpha, like noradrenaline, exposure of the cells increased the level of membrane G(i-alpha) proteins, it did not decrease but rather caused an increase in adenylyl cyclase responsiveness. This hypersensitivity may be due to concomitant alterations of other components, e.g., G-beta, G(s-alpha) and/or the cyclase itself, of this multi-subunit signal transduction system following TNF-alpha exposure.