Regulation of endothelial nitric-oxide synthase (NOS) S-glutathionylation by neuronal NOS: evidence of a functional interaction between myocardial constitutive NOS isoforms

Idigo, Winifred O;Reilly, Svetlana;Zhang, Mei Hua;Zhang, Yin Hua;Casadei, Barbara;et.al.
(2012) Journal of Biological Chemistry — Vol. 287, n° 52, p. 43665-43673 (2012)

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Authors
  • Idigo, Winifred O
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  • Reilly, Svetlana
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  • Zhang, Mei Hua
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  • Zhang, Yin Hua
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  • Balligand, Jean-LucUCLouvain
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  • Casadei, Barbara
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Abstract
Myocardial constitutive No production depends on the activity of both endothelial and neuronal NOS (eNOS and nNOS, respectively). Stimulation of myocardial β(3)-adrenergic receptor (β(3)-AR) produces a negative inotropic effect that is dependent on eNOS. We evaluated whether nNOS also plays a role in β(3)-AR signaling and found that the β(3)-AR-mediated reduction in cell shortening and [Ca(2+)](i) transient amplitude was abolished both in eNOS(-/-) and nNOS(-/-) left ventricular (LV) myocytes and in wild type LV myocytes after nNOS inhibition with S-methyl-l-thiocitrulline. LV superoxide (O(2)()) production was increased in nNOS(-/-) mice and reduced by l-N(ω)-nitroarginine methyl ester (l-NAME), indicating uncoupling of eNOS activity. eNOS S-glutathionylation and Ser-1177 phosphorylation were significantly increased in nNOS(-/-) myocytes, whereas myocardial tetrahydrobiopterin, eNOS Thr-495 phosphorylation, and arginase activity did not differ between genotypes. Although inhibitors of xanthine oxidoreductase (XOR) or NOX2 NADPH oxidase caused a similar reduction in myocardial O(2)(), only XOR inhibition reduced eNOS S-glutathionylation and Ser-1177 phosphorylation and restored both eNOS coupled activity and the negative inotropic and [Ca(2+)](i) transient response to β(3)-AR stimulation in nNOS(-/-) mice. In summary, our data show that increased O(2)() production by XOR selectively uncouples eNOS activity and abolishes the negative inotropic effect of β(3)-AR stimulation in nNOS(-/-) myocytes. These findings provide unequivocal evidence of a functional interaction between the myocardial constitutive NOS isoforms and indicate that aspects of the myocardial phenotype of nNOS(-/-) mice result from disruption of eNOS signaling.
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Idigo, W. O., Reilly, S., Zhang, M. H., Zhang, Y. H., Jayaram, R., Carnicer, R., Crabtree, M. J., Balligand, J.-L., & Casadei, B. (2012). Regulation of endothelial nitric-oxide synthase (NOS) S-glutathionylation by neuronal NOS: evidence of a functional interaction between myocardial constitutive NOS isoforms. Journal of Biological Chemistry, 287(52), 43665-43673. https://doi.org/10.1074/jbc.M112.412031 (Original work published 2012)