Activation of MTK1/MEKK4 by GADD45 through Induced N-C Dissociation and Dimerization-Mediated trans Autophosphorylation of the MTK1 Kinase Domain

Abstract

The mitogen-activated protein kinase (MAPK) module, composed of a MAPK, a MAPK kinase (MAPKK), and a MAPKK kinase (MAPKKK), is a cellular signaling device that is conserved throughout the eukaryotic world. In mammalian cells, various extracellular stresses activate two major subfamilies of MAPKs, namely, the Jun N-terminal kinases and the p38/stress-activated MAPK (SAPK). MTK1 (also called MEKK4) is a stress-responsive MAPKKK that is bound to and activated by the stress-inducible GADD45 family of proteins (GADD45␣/␤/␥). Here, we dissected the molecular mechanism of MTK1 activation by GADD45 proteins. The MTK1 N terminus bound to its C-terminal segment, thereby inhibiting the C-terminal kinase domain. This N-C interaction was disrupted by the binding of GADD45 to the MTK1 N-terminal GADD45-binding site. GADD45 binding also induced MTK1 dimerization via a dimerization domain containing a coiled-coil motif, which is essential for the trans autophosphorylation of MTK1 at Thr-1493 in the kinase activation loop. An MTK1 alanine substitution mutant at Thr-1493 has a severely reduced activity. Thus, we conclude that GADD45 binding induces MTK1 N-C dissociation, dimerization, and autophosphorylation at Thr-1493, leading to the activation of the kinase catalytic domain. Constitutively active MTK1 mutants induced the same events, but in the absence of GADD45.

Pregnane X Receptor PXR Activates the GADD45β Gene, Eliciting the p38 MAPK Signal and Cell Migration

“…This selection was made based on the fact that GADD45␤ is known to regulate phosphorylation of p38 MAPK (15,17,18) and the hypothesis that the gene needs to quickly respond to PXR activation to stimulate p38 MAPK phosphorylation within 30 min following RIF treatment. Increase of GADD45␤ mRNA was, in fact, detected as early as 15 min and peaked 60 min following RIF treatment in ShP51 cells but not in parent HepG2 cells (Fig.…”

“…Through protein-protein interactions with numerous signal molecules, GADD45␤ can regulate cellular signals for cell cycle, DNA repair, and apoptosis, depending on the types of stimuli and cells that are stimulated (14 -17). For example, GADD45␤ activates the p38 mitogen-activated protein kinase (MAPK) signal pathway via direct interaction with MTK1/MEKK4, a MAPK kinase kinase (15,17), repressing angiogenesis in the human pancreatic carcinoma Panc-1 cells (18).…”

Gadd45b Mediates Fas-induced Apoptosis by Enhancing the Interaction between p38 and Retinoblastoma Tumor Suppressor

“…Physical interaction of Gadd45 with MTK1 induces MTK1 N-to C-terminal dissociation, dimerization, and autophosphorylation and leads to the activation of the catalytic domain of the kinase (19,20). These studies have suggested that Gadd45 proteins may mediate activation of the p38/JNK MAPK pathway through MTK1 in response to environmental stress.…”

SAPK pathways and p53 cooperatively regulate PLK4 activity and centrosome integrity under stress

“…PLK4 did not bind to unactivated full-length MTK1, but it did so if GADD45b, a specific activator of MTK1 (ref. 32), was co-expressed (Fig. 1a, lanes 3 and 4).…”