Truncated IRAG variants modulate cGMP-mediated inhibition of human colonic smooth muscle cell contraction

Am J Physiol Cell Physiol. 2011 Dec;301(6):C1445-57. doi: 10.1152/ajpcell.00304.2010. Epub 2011 Aug 24.

Abstract

Nitric oxide (NO) induces relaxation of colonic smooth muscle cells predominantly by cGMP/cGMP-dependent protein kinase I (cGKI)-induced phosphorylation of the inositol 1,4,5-trisphosphate receptor (IP(3)R)-associated cGMP kinase substrate (IRAG), to block store-dependent calcium signaling. In the present study we analyzed the structure and function of the human IRAG/MRVI1 gene. We describe four unique first exon variants transcribed from individual promoters in diverse human tissues. Tissue-specific alternative splicing with exon skipping and alternative splice donor and acceptor site usage further increases diversity of IRAG mRNA variants that encode for NH(2)- and COOH-terminally truncated proteins. At the functional level, COOH-terminally truncated IRAG variants lacking both the cGKI phosphorylation and the IP(3)RI interaction site counteract cGMP-mediated inhibition of calcium transients and relaxation of human colonic smooth muscle cells. Since COOH-terminally truncated IRAG mRNA isoforms are widely expressed in human tissues, our results point to an important role of IRAG variants as negative modulators of nitric oxide/cGKI-dependent signaling. The complexity of alternative splicing of the IRAG gene impressively demonstrates how posttranscriptional processing generates functionally distinct proteins from a single gene.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Blotting, Western
  • Cell Line
  • Colon / metabolism*
  • Cyclic GMP / metabolism
  • Cyclic GMP-Dependent Protein Kinases / metabolism
  • Gene Expression Profiling*
  • Humans
  • Immunoprecipitation
  • Membrane Proteins / genetics*
  • Membrane Proteins / metabolism
  • Molecular Sequence Data
  • Muscle Contraction / genetics*
  • Muscle, Smooth / metabolism*
  • Phosphoproteins / genetics*
  • Phosphoproteins / metabolism
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • RNA Isoforms / genetics
  • RNA Isoforms / metabolism
  • RNA Processing, Post-Transcriptional
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transfection

Substances

  • MRVI1 protein, human
  • Membrane Proteins
  • Phosphoproteins
  • Protein Isoforms
  • RNA Isoforms
  • Cyclic GMP-Dependent Protein Kinases
  • Cyclic GMP