L-type Ca(2+) channel activation regulates induction of c-fos transcription by hypoxia.

Abstract

In the present study we examined the intracellular pathways that link hypoxia to activation of c-fos gene expression. Experiments were performed on rat pheocromocytoma-12 (PC-12) cells. c-fos mRNA and promoter activities were analyzed by RT-PCR and reporter gene assays, respectively. BAPTA, a Ca(2+) chelator, inhibited c-fos mRNA and promoter activation by hypoxia. Nitrendipine, an L-type Ca(2+)-channel blocker, abolished, whereas BAY K 8644, an L-type channel agonist, enhanced c-fos activation by hypoxia. Ca(2+) currents were augmented reversibly by hypoxia, suggesting that Ca(2+) influx mediated by L-type Ca(2+) channels is essential for c-fos activation by hypoxia. We next determined downstream pathways activated by intracellular Ca(2+) concentration. Immunoblot analysis revealed Ca(2+)/calmodulin-dependent kinase II (CaMKII) protein in PC-12 cells and revealed that hypoxia increased the enzyme activity. KN-93, a CaMK inhibitor, blocked CaMKII activation and c-fos promoter stimulation by hypoxia. Ectopic expression of an active mutant of CaMKII (pCaMKII290) stimulated c-fos promoter activity under normoxia. Hypoxia increased phosphorylation of CREB at the serine residue 133 (Ser-133), and KN-93 attenuated this effect. Point mutations at the Ca(2+)/cAMP-responsive cis-element (Ca/CRE) attenuated, whereas point mutations in the serum-responsive cis-element (SRE) abolished transcriptional activation of c-fos by hypoxia. These results demonstrate that c-fos activation by hypoxia involves CaMK activation and CREB phosphorylation at Ser-133 and requires Ca/CRE and SRE. These observations demonstrate that Ca(2+)-dependent signaling pathways play a crucial role in induction of c-fos gene expression, which may underlie long-term adaptive responses to hypoxia.

• Authors: D R Premkumar, R R Mishra, J L Overholt, M S Simonson, N S Cherniack, N R Prabhakar
• Journal: Journal of applied physiology (Bethesda, Md. : 1985) (J Appl Physiol (1985)) Vol. 88 Issue 5 Pg. 1898-906 (May 2000) ISSN: 8750-7587 [Print] United States
• PMID: 10797155 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Calcium Channel Agonists
• Calcium Channel Blockers
• Calcium Channels, L-Type
• Chelating Agents
• Cyclic AMP Response Element-Binding Protein
• Enzyme Inhibitors
• Proto-Oncogene Proteins c-fos
• RNA, Messenger
• Egtazic Acid
• Calcium-Calmodulin-Dependent Protein Kinase Type 2
• Calcium-Calmodulin-Dependent Protein Kinases
• 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid

Topics

• Animals
• Calcium Channel Agonists (pharmacology)
• Calcium Channel Blockers (pharmacology)
• Calcium Channels, L-Type (physiology)
• Calcium-Calmodulin-Dependent Protein Kinase Type 2
• Calcium-Calmodulin-Dependent Protein Kinases (antagonists & inhibitors)
• Chelating Agents (pharmacology)
• Cyclic AMP Response Element-Binding Protein (metabolism)
• Egtazic Acid (analogs & derivatives, pharmacology)
• Enzyme Inhibitors (pharmacology)
• Gene Expression Regulation (physiology)
• Hypoxia (genetics, physiopathology)
• PC12 Cells
• Phosphorylation
• Point Mutation (physiology)
• Promoter Regions, Genetic (physiology)
• Proto-Oncogene Proteins c-fos (genetics)
• RNA, Messenger (metabolism)
• Rats
• Transcription, Genetic (physiology)