Bromelain, from pineapple stems, proteolytically blocks activation of extracellular regulated kinase-2 in T cells

Recently, it has emerged that extracellular proteases have specific regulat ory roles in modulating immune responses. Proteases may act as signaling mo lecules to activate the Raf-1/extracellular regulated kinase (ERK)-2 pathwa y to participate in mitogenesis, apoptosis, and cytokine production. Most r eports on the role of protease-mediated cell signaling, however, focus on t heir stimulatory effects, In this study, we show for the first time that ex tracellular proteases may also block signal transduction. We show that brom elain, a mixture of cysteine proteases from pineapple stems, blocks activat ion of ERK-2 in Th0 cells stimulated via the TCR with anti-CD3 epsilon mAb, or stimulated with combined PMA and calcium ionophore, The inhibitory acti vity of bromelain was dependent on its proteolytic activity, as ERK-2 inhib ition was abrogated by E-64, a selective cysteine protease inhibitor. Howev er, inhibitory effects were not caused by nonspecific proteolysis, as the p rotease trypsin had no effect on ERK activation. Bromelain also inhibited P MA-induced IL-2, IFN-gamma, and IL-4 mRNA accumulation, but had no effect o n TCR-induced cytokine mRNA production. This data suggests a critical requi rement for ERK-2 in PMA-induced cytokine production, but nut TCR-induced cy tokine production. Bromelain did not act on ERK-2 directly, as it also inhi bited p21(ras) activation, an effector molecule upstream from ERK-2 in the Raf-1/MEK/ERK-2 kinase signaling cascade. The results indicate that bromela in is a novel inhibitor of T cell signal transduction and suggests a novel role for extracellular proteases as inhibitors of intracellular signal tran sduction pathways.