IRAG determines nitric oxide- and atrial natriuretic peptide-mediated smooth muscle relaxation.
OBJECTIVE
Nitric oxide (NO) and atrial natriuretic peptide (ANP) signalling via cGMP controls smooth muscle tone. One important signalling pathway of cGMP-dependent protein kinase type I (cGKI) is mediated by IRAG (IP(3) receptor associated cGKI substrate) which is highly expressed in smooth muscle tissues. To elucidate the role of IRAG for NO- and ANP-mediated smooth muscle tone regulation, cGKI localization, and for its possible function in blood pressure adjustment, we generated IRAG-knockout mice by targeted deletion of exon 3.


RESULTS
IRAG deletion prevented stable interaction of IP(3) receptor type I (IP(3)RI) with cGKIbeta determined by cGMP affinity chromatography. Confocal microscopy in vascular smooth muscle cells (VSMCs) showed that localization of cGKIbeta and cGKIalpha did not change in absence of IRAG. NO-, ANP-, and cGMP-dependent relaxation of hormone-contracted aortic vessels and colon was significantly affected in IRAG-knockout mice. The suppression of cGMP-induced relaxation was not rescued by selective expression of cGKIbeta in smooth muscle from cGKIbeta-transgenic mice. NO-, ANP-, and cGMP-mediated inhibition of the hormone-induced increase in intracellular calcium concentration measured by Fura2 was suppressed in IRAG-deficient VSMC. Telemetric measurements revealed that IRAG-deficient animals exhibited normal basal tone, but were resistant to blood pressure reduction induced by lipopolysaccharide-treatment.


CONCLUSIONS
These findings indicate that signalling of cGKIbeta via IRAG is an essential functional part for regulation of smooth muscle tone and of intracellular calcium by NO (exogenously applicated or endogenously synthesized) and by ANP. IRAG signalling does not modulate basal tone but might be important for blood pressure regulation under pathophysiological conditions.
Desch
Matthias
M
Pharmakologie und Toxikologie, Universität Regensburg, Universitätstr. 31, D-93055 Regensburg, Germany.
Sigl
Katja
K
Hieke
Bernhard
B
Salb
Katharina
K
Kees
Frieder
F
Bernhard
Dominik
D
Jochim
Angela
A
Spiessberger
Beate
B
Höcherl
Klaus
K
Feil
Robert
R
Feil
Susanne
S
Lukowski
Robert
R
Wegener
Jörg W
JW
Hofmann
Franz
F
Schlossmann
Jens
J
D000818:Animals; D001011:Aorta / Q000378:metabolism; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D001794:Blood Pressure; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity; D003106:Colon / Q000378:metabolism; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D005091:Exons; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D009126:Muscle Relaxation; D009130:Muscle, Smooth / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D010750:Phosphoproteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D014664:Vasodilation
D053496:Inositol 1,4,5-Trisphosphate Receptors; D047908:Intracellular Signaling Peptides and Proteins; C406072:MRVI1 protein, Bos taurus; D008565:Membrane Proteins; C485785:Mrvi1 protein, mouse; D010750:Phosphoproteins; D009569:Nitric Oxide; D009320:Atrial Natriuretic Factor; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases; C569720:Prkg1 protein, mouse; D002118:Calcium