c-Jun NH2-terminal kinase 2α2 promotes the tumorigenicity of human glioblastoma cells

c-Jun NH2-terminal kinases (JNK) are members of the mitogen-activated protein kinase family and have been implicated in the formation of several human tumors, especially gliomas. We have previously shown that a 55 kDa JNK isoform is constitutively active in 86% of human brain tumors and then showed that it is specifically a JNK2 isoform and likely to be either JNK2 alpha 2 or JNK2 beta 2. Notably, we found that only JNK2 isoforms possess intrinsic autophosphorylation activity and that JNK2 alpha 2 has the strongest activity. In the present study, we have further explored the contribution of JNK2 isoforms to brain tumor formation. Analysis of mRNA expression by reverse transcription-PCR revealed that JNK2 alpha 2 is expressed in 91% (10 of 11) of glioblastoma tumors, whereas JNK2 beta 2 is found in only 27% (3 of 11) of tumors. Both JNK2 alpha 2 and JNK2 beta 2 mRNAs are expressed in normal brain (3 of 3). Using an antibody specific for JNK2 alpha isoforms, we verified that JNK2 alpha 2 protein is expressed in 88.2% (15 of 17) of glioblastomas, but, interestingly, no JNK2 alpha 2 protein was found in six normal brain samples. To evaluate biological function, we transfected U87MG cells with green fluorescent protein-tagged versions of JNK1 alpha 1, JNK2 alpha 2, and JNK2 alpha 2APF (a dominant-negative mutant), and derived cell fines with stable expression. Each cell fine was evaluated for various tumorigenic variables including cellular growth, soft agar colony formation, and tumor formation in athymic nude mice. In each assay, JNK2 alpha 2 was found to be the most effective in promoting that phenotype. To identify effectors specifically affected by JNK2 alpha 2, we analyzed gene expression. Gene profiling showed several genes whose expression was specifically up-regulated by JNK2 alpha 2 but down-regulated by JNK2 alpha 2APF, among which eukaryotic translation initiation factor 4E (eIF4E) shows the greatest change. Because AKT acts on eIF4E, we also examined AKT activation. Unexpectedly, we found that JNK2 alpha 2 could specifically activate AKT. Our data provides evidence that JNK2 alpha 2 is the major active JNK isoform and is involved in the promotion of proliferation and growth of human glioblastoma tumors through specific activation of AKT and overexpression of eIF4E.