viacount

Thapsigargin induces expression of ATF3 in human keratinocytes involving Ca2+ ions and c-Jun N-terminal protein kinase.

Thapsigargin is a specific inhibitor of the SERCA ATPase of the endoplasmic reticulum. Here, we show that stimulation of human HaCaT keratinocytes with nanomolar concentrations of thapsigargin triggers expression of ATF3, a basic-region leucin zipper transcription factor. ATF3 expression was also upregulated in thapsigargin-stimulated glioma cells, hepatoma cells, retinal pigment epithelial cells, and airway epithelial cells. Thapsigargin-induced upregulation of ATF3 expression in keratinocytes was attenuated by BAPTA-AM, or by expression of the Ca(2+)-binding protein parvalbumin in the cytosol of HaCaT cells, but not by a panel of pharmacological agents that chelate extracellular Ca(2+) (EGTA) or inhibit either ryanodine receptors (dantrolene) or voltage-gated Ca(2+) channels (nifedipine). Hence, elevated levels of intracellular Ca(2+), released from intracellular stores, are essential for the effect of thapsigargin on the biosynthesis of ATF3. The thapsigargin-induced signaling pathway was blocked by expression of either MAP kinase phophatases-1 or -5. Experiments involving pharmacological and genetic tools revealed the importance of c-Jun N-terminal protein kinase (JNK) within the signaling cascade, while inhibition of extracellular signal-regulated protein kinase or p38 protein kinase did not attenuate thapsigargin-induced expression of ATF3. Functional studies showed that treatment of HaCaT keratinocytes with thapsigargin led to a 2-fold induction of caspase-3/7 activity. The upregulation of caspase-3/7 activity in thapsigargin-stimulated HaCaT cells was attenuated by inhibition of JNK. Together, these data show that stimulation of HaCaT cells with thapsigargin induces a specific signaling pathway in keratinocytes involving activation of JNK, biosynthesis of ATF3, and upregulation of caspase-3/7 activity.

Document Type:

Reference

Product Catalog Number:

06-659

Product Catalog Name:

Anti-phospho-c-Jun (Ser73) Antibody