Activation of ERK1/2 and TNF-α production are mediated by calcium/ calmodulin, and PKA signaling pathways during Mycobacterium bovis infection

Mycobacterium bovis bacillus Calmette-Guérin (BCG)-induced tumor necrosis factor (TNF)-α secretion via an extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase-dependent mechanism is an important host defence mechanism against Mycobacterium tuberculosis in human monocytes. We now define distinct signaling pathways that regulate induction of TNF-α and activation of ERK1/2 by intracellular signaling mechanisms during M. bovis infection. We determined that M. bovis BCG-induced ERK 1/2 activation occurs through a mechanism that requires intracellular calcium and likely involves a calmodulin-sensitive step. In contrast, M. bovis BCG can induce p38 mapk activation by a calcium (Ca²⁺)/calmodulin-independent mechanism. Interestingly, we present evidence that M. bovis BCG activates protein kinase A (PKA), since pretreatment of monocytes with H-89, a inhibitor of PKA activity, blocked the ability of M. bovis BCG to induce ERK1/2 activation. These results were further supported by the fact that treatment of cells with KT5720, another well-described inhibitor of PKA activity, significantly diminished the effect of M. bovis BCG on ERK1/2 activation. Furthermore, secretion of TNF-α in M. bovis-infected human monocytes was also dependent on Ca²⁺/ calmodulin, and PKA pathways. Finally, addition of H-89 significantly diminished TNF-α mRNA expression in M. bovis-infected human monocytes. These results indicate that the Ca²⁺/calmodulin, and PKA pathways play important regulatory roles in monocyte signaling upon M. bovis infection.