Thapsigargin induces expression of activating transcription factor 3 in human keratinocytes involving Ca2+ ions and c-Jun N-terminal protein kinase

• Published in: Molecular pharmacology Vol. 78; no. 5; pp. 865 - 876
• Main Authors: Spohn, Daniel, Rössler, Oliver G, Philipp, Stephan E, Raubuch, Michael, Kitajima, Shigetaka, Griesemer, Désirée, Hoth, Markus, Thiel, Gerald
• Format: Journal Article
• Language: English
• Published: United States 01.11.2010
• Subjects: Activating Transcription Factor 1 - biosynthesis; Anisomycin - pharmacology; Apoptosis - drug effects; Calcium - metabolism; Calcium-Transporting ATPases - antagonists & inhibitors; Caspase 3 - metabolism; Caspase 7 - metabolism; Cations, Divalent; Cell Line; Dual Specificity Phosphatase 1 - metabolism; Dual-Specificity Phosphatases - metabolism; Enzyme Activation; Humans; JNK Mitogen-Activated Protein Kinases - metabolism; Keratinocytes - cytology; Keratinocytes - drug effects; Keratinocytes - metabolism; Mitogen-Activated Protein Kinase Phosphatases - metabolism; p38 Mitogen-Activated Protein Kinases - physiology; Signal Transduction; Thapsigargin - pharmacology; Up-Regulation
• ISSN: 1521-0111
• DOI: 10.1124/mol.110.067637

Thapsigargin is a specific inhibitor of the sarco/endoplasmic reticulum Ca(2+) ATPase of the endoplasmic reticulum. Here, we show that stimulation of human HaCaT keratinocytes with nanomolar concentrations of thapsigargin triggers expression of activating transcription factor (ATF) 3, a basic-region leucin zipper transcription factor. ATF3 expression was also up-regulated in thapsigargin-stimulated glioma cells, hepatoma cells, retinal pigment epithelial cells, and airway epithelial cells. Thapsigargin-induced up-regulation of ATF3 expression in keratinocytes was attenuated by BAPTA-acetoxymethyl ester 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 mitogen-activated protein kinase phosphatase-1 or -5. Experiments involving pharmacological and genetic tools revealed the importance of c-Jun N-terminal protein kinase (JNK) within the signaling cascade, whereas 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 up-regulation 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 up-regulation of caspase-3/7 activity.