Hyperexpression of c-jun and c-fos mRNA's are cndpoints of the p21ras-dependent pathway of transformation by p210 BCR-ABL, the Philadelphia chromosome product. We studied the contribution of JNK, the cjun N-terminal kinase that augments, by phosphorylation, c-jun/AP-1-dependent transact!vation of c-jun transcription and elk-1 -dependent c-fos transcription, to explain c-jun and c-fos hyperexpression. By titration of cell extracts in a kinase assay involving GST-c-jun (5-89) as substrate, there was at least 6-fold greater activity of JNK within p210 BCR-ABL transformed cells compared to IL-3 treated parental cells. Corresponding to heightened JNK activity downstream of p210 BCR-ABL, there was a unique transactivating complex (AP-1 complex 2} consisting of c-jun and c-fos, binding to the proximal jun promoter (jun 1) TRE in gel mobility shifts. By contrast, the AP-1 contained within IL-3-dependent H7 parental cells was exclusively c-jun/ATF-2 heterodimer (complex 1). 5'-cjun mediated transcriptional activity in BCR-ABL transformed cells was totally dependent on binding of AP-1 by junl TRE, as demonstrated in reporter assays after site-directed mutagenesis of jun 1 TRE. Dose-dependent inhibition of JNK activity within BCR-ABL transformed cells resulted from treatment with cyclosporin A (CsA), an inhibitor of the Ca2+-calmodulin dependent enzyme, calcineurin (CN) [also called PP2B], or by treatment with W7, a calmodulin inhibitor. Dose-dependent inhibition of c-jun transcript expression on Northern blots followed JNK inhibition by CsA. Inhibition of c-jun mRNA by cyclosporin was a unique property of BCR-ABL transformed cells (absent in parental cells) and CsA treatment led to depletion of the unique c-jun/c-fos AP-1 complex (complex 2) binding to the 5' jun 1 site. Thus, BCR-ABL transformation requires JNK activity for c-jun and c-fos expression and a Ca2+-dependent JNK pathway unique to transformed cells is operating.
|Original language||English (US)|
|Number of pages||1|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology
- Cancer Research