Destabilization of TNF-alpha mRNA by Rapamycin

Abstract

Stimulation of mast cells through the high affinity IgE receptor (Fc epsilon RI) induces degranulation, lipid mediator release, and cytokine secretion leading to allergic reactions. Although various signaling pathways have been characterized to be involved in the Fc epsilon RI-mediated responses, little is known about the precious mechanism for the expression of tumor necrosis factor-alpha (TNF-alpha) in mast cells. Here, we report that rapamycin, a specific inhibitor of mammalian target of rapamycin (mTOR), reduces the expression of TNF-alpha in rat basophilic leukemia (RBL-2H3) cells. IgE or specific antigen stimulation of RBL-2H3 cells increases the expression of INF-alpha and activates various signaling molecules including S6K1, Akt and p38 MAPK. Rapamycin specifically inhibits antigen-induced INF-alpha mRNA level, while other kinase inhibitors have no effect on TNF-alpha mRNA level. These data indicate that mTOR signaling pathway is the main regulation mechanism for antigen-induced TNF-alpha expression. TNF-alpha mRNA stability analysis using reporter construct containing TNF-alpha adenylate/uridylate-rich elements (AREs) shows that rapamycin destabilizes TNF-alpha mRNA via regulating the AU-rich element of TNF-alpha mRNA. The antigen-induced activation of S6K1 is inhibited by specific kinase inhibitors including mTOR, PI3K, PKC and Ca(2+)chelator inhibitor, while TNF-alpha mRNA level is reduced only by rapamycin treatment. These data suggest that the effects of rapamycin on the expression of TNF-alpha mRNA are not mediated by S6K1 but regulated by mTOR. Taken together, our results reveal that mTOR signaling pathway is a novel regulation mechanism for antigen-induced TNF-alpha expression in RBL-2H3 cells.