The effect of tyrosine nitration of L-type Ca2+ channels on excitation-transcription coupling in colonic inflammation.

Abstract

BACKGROUND AND PURPOSE:

Excitation-transcriptional coupling involves communication between plasma membrane ion channels and gene expression in the nucleus. Calcium influx through L-type Ca(2+) channels induces phosphorylation of the transcription factor, cyclic-AMP response element binding protein (CREB) and downstream activation of the cyclic-AMP response element (CRE) promoter regions. Tyrosine nitration of Ca(2+) channels attenuates interactions with c-Src kinase, decreasing Ca(2+) channel currents and smooth muscle contraction during colonic inflammation. In this study we examined the effect of tyrosine nitration and colonic inflammation on Ca(2+) channel mediated phosphorylation of CREB and CRE activation.

EXPERIMENTAL APPROACH:

CREB and phospho-CREB were detected by Western blots and CRE activation measured by dual luciferase assay. Chinese hamster ovary (CHO) cells were transfected with hCa(v)1.2b and hCa(v)1.2b c-terminal mutants. Colonic inflammation was induced by intracolonic instillation of 2,4,6 trinitrobenzene sulphonic acid in mouse colon.

KEY RESULTS:

In hCa(v)1.2b transfected CHO cells and in native colonic smooth muscle, depolarization with 80 mM KCl induced CREB phosphorylation (pCREB). Treatment with peroxynitrite inhibited KCl-induced pCREB. Following experimental colitis, KCl-induced CREB phosphorylation was abolished in smooth muscle, concomitant with tyrosine nitration of Ca(2+) channels. Depolarization increased CRE activation in hCa(v)1.2b CHO cells by 2.35 fold which was blocked by nifedipine and by protein nitration of Ca(2+) channels with peroxynitrite. The Src-kinase inhibitor, PP2, blocked depolarization-induced CRE activation. Mutation of the C-terminus tyrosine residue, Y2134F, but not Y1861F, blocked CRE activation.

CONCLUSIONS AND IMPLICATIONS:

Post-translational modification of Ca(2+) channels due to tyrosine nitration modified excitation-transcriptional coupling in colonic inflammation.

• Authors: M Kang, G R Ross, H I Akbarali
• Journal: British journal of pharmacology (Br J Pharmacol) Vol. 159 Issue 6 Pg. 1226-35 (Mar 2010) ISSN: 1476-5381 [Electronic] England
• PMID: 20128810 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)

Chemical References

• Calcium Channels, L-Type
• Cyclic AMP Response Element-Binding Protein
• Protein Kinase Inhibitors
• Peroxynitrous Acid
• Tyrosine
• src-Family Kinases

Topics

• Animals
• Blotting, Western
• CHO Cells
• Calcium Channels, L-Type (genetics, metabolism)
• Colitis (metabolism)
• Cricetinae
• Cricetulus
• Cyclic AMP Response Element-Binding Protein (genetics, metabolism)
• Disease Models, Animal
• Excitation Contraction Coupling (drug effects)
• Humans
• Immunohistochemistry
• Immunoprecipitation
• Male
• Membrane Potentials (drug effects)
• Mice
• Mice, Inbred BALB C
• Muscle, Smooth (drug effects, metabolism)
• Patch-Clamp Techniques
• Peroxynitrous Acid (pharmacology)
• Protein Kinase Inhibitors (pharmacology)
• Response Elements (genetics)
• Reverse Transcriptase Polymerase Chain Reaction
• Transcriptional Activation (drug effects)
• Transfection
• Tyrosine (metabolism)
• src-Family Kinases (antagonists & inhibitors)