Constitutive Activity of JNK2α2 Is Dependent on a Unique Mechanism of MAPK Activation

Abstract

c-Jun N-terminal kinases (JNKs) are part of the mitogen-activated protein kinase (MAPK) family and are important regulators of cell growth, proliferation, and apoptosis. Typically, a sequential series of events are necessary for MAPK activation: phosphorylation, dimerization, and then subsequent translocation to the nucleus. Interestingly, a constitutively active JNK isoform, JNK2␣2, possesses the ability to autophosphorylate and has been implicated in several human tumors, including glioblastoma multiforme. Because overexpression of JNK2␣2 enhances several tumorigenic phenotypes, including cell growth and tumor formation in mice, we studied the mechanisms of JNK2␣2 autophosphorylation and autoactivation. We find that JNK2␣2 dimerization in vitro and in vivo occurs independently of its autophosphorylation but is dependent on nine amino acids, known as the ␣-region. Alanine scanning mutagenesis of the ␣-region reveals that five specific mutants (L218A, K220A, G221A, I224A, and F225A) prevent JNK2␣2 dimerization rendering JNK2␣2 inactive and incapable of stimulating tumor formation. Previous studies coupled with additional mutagenesis of neighboring isoleucines and leucines (I208A, I214A, I231A, and I238A) suggest that a leucine zipper may play an important role in JNK2␣2 homodimerization. We also show that a kinase-inactive JNK2␣2 mutant can interact with and inhibit wild type JNK2␣2 autophosphorylation, suggesting that JNK2␣2 undergoes trans-autophosphorylation. Together, our results demonstrate that JNK2␣2 differs from other MAPK proteins in two major ways; its autoactivation/autophosphorylation is dependent on dimerization, and dimerization most likely precedes autophosphorylation. In addition, we show that dimerization is essential for JNK2␣2 activity and that prevention of dimerization may decrease JNK2␣2 induced tumorigenic phenotypes.

Molecular cloning and characterization of novel human JNK2 (MAPK9) transcript variants that show different stimulation activities on AP-1

“…The α and β regions determine the binding activities of JNK2 substrates (5,6), and the α region is also important for JNK2 in autophosphorylation (15) and dimerization (17). Consistently with this view, JNK2α3 showed higher cis-AP-1 and different substrate activities compared with JNK2β3, further suggesting that the α and β regions are directly involved in the divergent function of various JNK2 isoforms.…”

Thr-370 Is Responsible for CDK11p58 Autophosphorylation, Dimerization, and Kinase Activity

“…Moreover, autophosphorylation/autoactivation of protein kinase is usually related to the homodimerization. For example, disruption of JNK2␣2 dimerization through specific mutations in the ␣-region resulted in loss of nuclear localization, loss of autophosphorylation, and decreased tumorigenicity (18). Thus, the autophosphorylation of CDK11 p58 may be related to its dimerization.…”

“…Protein kinase autophosphorylation of activation segment residues is a common regulatory mechanism in phosphorylation-dependent signaling cascades. For example, the c-Jun N-terminal kinase (JNK), which belongs to the mitogen-activated protein kinase (MAPK) family, can be autophosphorylated/autoactivated, which is a consequence following homodimerization of the kinase (18). MTK1 (also called MEKK4) is a stress-responsive MAPKKK.…”

MicroRNA-206 Regulates Cell Movements during Zebrafish Gastrulation by Targeting prickle1a and Regulating c-Jun N-Terminal Kinase 2 Phosphorylation

“…5G). To investigate whether the decrease of JNK2 phosphorylation was responsible for the CE defects in the mir206-misexpressed embryos, 50 pg human JNK2␣2 mRNA, which encodes a constitutively active JNK2 isoform (11,41), was coinjected with mir206 precursors or 206-MOs. The result showed that the CE defects indicated by the expression pattern of ntl were rescued in about 75% mir206-misexpressed embryos (Fig.…”