Depolarization-induced differentiation of PC12 cells is mediated by phospholipase D2 through the transcription factor CREB pathway.

Abstract

The present study examined the role of phospholipase D2 (PLD2) in the regulation of depolarization-induced neurite outgrowth and the expression of growth-associated protein-43 (GAP-43) and synapsin I in rat pheochromocytoma (PC12) cells. Depolarization of PC12 cells with 50 mmol/L KCl increased neurite outgrowth and elevated mRNA and protein expression of GAP-43 and synapsin I. These increases were suppressed by inhibition of Ca2+-calmodulin-dependent protein kinase II (CaMKII), PLD, or mitogen-activated protein kinase kinase (MEK). Knockdown of PLD2 by small interfering RNA (siRNA) suppressed the depolarization-induced neurite outgrowth, and the increase in GAP-43 and synapsin I expression. Depolarization evoked a Ca2+ rise that activated various signaling enzymes and the cAMP response element-binding protein (CREB). Silencing CaMKIIdelta by siRNA blocked KCl-induced phosphorylation of proline-rich protein tyrosine kinase 2 (Pyk2), Src kinase, and extracellular signal-regulated kinase (ERK). Inhibition of Src or MEK abolished phosphorylation of ERK and CREB. Furthermore, phosphorylation of Pyk2, ERK, and CREB was suppressed by the PLD inhibitor, 1-butanol and transfection of PLD2 siRNA, whereas it was enhanced by over-expression of wild-type PLD2. Depolarization-induced PLD2 activation was suppressed by CaMKII and Src inhibitors, but not by MEK or protein kinase A inhibitors. These results suggest that the signaling pathway of depolarization-induced PLD2 activation was downstream of CaMKIIdelta and Src, and upstream of Pyk2(Y881) and ERK/CREB, but independent of the protein kinase A. This is the first demonstration that PLD2 activation is involved in GAP-43 and synapsin I expression during depolarization-induced neuronal differentiation in PC12 cells.

• Authors: Yoshiko Banno, Satoshi Nemoto, Masashi Murakami, Masashi Kimura, Yoshihito Ueno, Kenji Ohguchi, Akira Hara, Yukio Okano, Yukio Kitade, Minoru Onozuka, Takashi Murate, Yoshinori Nozawa
• Journal: Journal of neurochemistry (J Neurochem) Vol. 104 Issue 5 Pg. 1372-86 (Mar 2008) ISSN: 1471-4159 [Electronic] England
• PMID: 18004999 (Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Cyclic AMP Response Element-Binding Protein
• Enzyme Inhibitors
• GAP-43 Protein
• Neuromuscular Depolarizing Agents
• Synapsins
• Transcription Factors
• Calcium-Calmodulin-Dependent Protein Kinase Type 2
• phospholipase D2
• Phospholipase D

Topics

• Animals
• Calcium-Calmodulin-Dependent Protein Kinase Type 2 (antagonists & inhibitors, metabolism)
• Cell Differentiation (drug effects, genetics, physiology)
• Cyclic AMP Response Element-Binding Protein (physiology)
• Enzyme Inhibitors (pharmacology)
• GAP-43 Protein (biosynthesis, genetics)
• Neurites (drug effects, enzymology, metabolism)
• Neuromuscular Depolarizing Agents (pharmacology)
• PC12 Cells
• Phospholipase D (antagonists & inhibitors, biosynthesis, genetics, physiology)
• Rats
• Signal Transduction (drug effects, genetics, physiology)
• Synapsins (biosynthesis, genetics)
• Transcription Factors (physiology)