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Properties of NO‐activated guanylyl cyclases expressed in cells

2003; Wiley; Volume: 139; Issue: 5 Linguagem: Inglês

10.1038/sj.bjp.0705318

1476-5381

Barry Gibb, Victoria Wykes, John Garthwaite,

Adenosine and Purinergic Signaling

Physiological nitric oxide (NO) signal transduction occurs through activation of guanylyl cyclase (GC)‐coupled receptors, resulting in cGMP accumulation. There are five possible receptors: four heterodimers ( α 1 β 1, α 2 β 1, α 1 β 2, α 2 β 2) and a presumed homodimer ( νβ 2). The present study investigated the kinetic and pharmacological properties of all these putative receptors expressed in COS‐7 (or HeLa) cells. All exhibited NO‐activated GC activity, that of α 1 β 1 and α 2 β 1 being much higher than that of the β 2‐containing heterodimers or νβ 2. All were highly sensitive NO detectors. Using clamped NO concentrations, EC 50 values were 1 n M for α 1 β 1 and 2 n M for α 2 β 1. With α 1 β 2, α 2 β 2 and νβ 2, the EC 50 was estimated to be lower, about 8 n M . All the GCs displayed a marked desensitising profile of activity. Consistent with this property, the concentration–response curves were bell‐shaped, particularly those of the β 2 heterodimers and νβ 2. Confocal microscopy of cells transfected with the fluorescently tagged β 2 subunit suggested targeting to the endoplasmic reticulum through its isoprenylation sequence, but no associated particulate GC activity was detected. The NO‐stimulated GC activity of all heterodimers and νβ 2 was inhibited by 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one and, except for νβ 2, was enhanced by the allosteric activator YC‐1. It is concluded that all the four possible heterodimers, as well as the putative νβ 2 homodimer, can function as high‐affinity GC‐coupled NO receptors when expressed in cells. They exhibit differences in NO potency, maximal GC activity, desensitisation kinetics and possibly subcellular location but, except for νβ 2, cannot be differentiated using existing pharmacological agents. British Journal of Pharmacology (2003) 139 , 1032–1040. doi: 10.1038/sj.bjp.0705318