Mechanisms regulating the function of oncoprotein kinase Tpl2 in macrophages and HTLV-I-transformed T cells

Macrophages express a family of pattern recognition receptors called Toll-like receptors which in response to stimulation activate various downstream signaling pathways leading to induction of genes encoding inflammatory mediators. Tpl2, a MAP3K in the ERK signaling pathway, plays a critical role in regulating macrophage activation by Toll-like receptor agonists. While the role of p105 and IKK in Tpl2 stabilization and activation have been established, exact mechanisms regulating Tpl2 function remain poorly understood. Results presented here show that p105 functions as a substrate and competitive inhibitor of Tpl2 and when bound to p105, Tpl2 is restricted from gaining access to its substrate MEK1. However, p105-associated Tpl2 remains kinetically active and uses p105 as intra-complex substrate. Tpl2 directly phosphorylates p105 in its C-terminal domain, at a site distinct from the IKKβ recognition motif. In addition, the long (Tpl2L) and short (Tpl2S) isoforms of Tpl2 show marked differences in their rate of release from p105 and subsequent degradation. Compared to Tpl2S, Tpl2L is much more rapidly released and degraded in response to signal stimulation. Interestingly, such a rapid response of Tpl2L is associated with its phosphorylation which is likely mediated by IKKβ. The N-terminal unique region of Tpl2 L contains a phosphorylation site for IKKβ and additional regulatory sequences, which appear to regulate its rapid release from p105 and degradation.;Evidence is also presented to show aberrant activation of Tpl2 in HTLV-I transformed T cells. HTLV-I induced T-cell transformation is largely mediated by the viral gene-encoded protein called Tax. Tax is known to activate IKK and its downstream transcription factor NF-κB, yet NF-κB activation is not sufficient for inducing T-cell transformation. Results presented here show that Tax also mediates activation of the oncoprotein kinase Tpl2. Constitutive activation of Tpl2 is seen in a large panel of HTLV-I-infected T cell lines. While Tpl2 is tightly regulated by p105 in normal cells, in HTLV-I-transformed T cells Tpl2 is predominantly dissociated from p105 and remains constitutively active. Interestingly, free Tpl2 is also remarkably stable. Activated Tpl2 in HTLV-I-transformed T cells not only contributes to MEK1 activation but also synergizes with Tax in activating NF-κB.