Abstract
Soluble guanylyl cyclase (sGC), a ubiquitously expressed heme-containing receptor for nitric oxide (NO), is a key mediator of NO-dependent processes. In addition to NO, a number of synthetic compounds that target the heme-binding region of sGC and activate it in a NO-independent fashion have been described. We report here that dicyanocobinamide (CN2-Cbi), a naturally occurring intermediate of vitamin B12 synthesis, acts as a sGC coactivator both in vitro and in intact cells. Heme depletion or heme oxidation does not affect CN2-Cbi-dependent activation. Deletion mutagenesis demonstrates that CN2-Cbi targets a new regulatory site and functions though a novel mechanism of sGC activation. Unlike all known sGC regulators that target the N-terminal regulatory regions, CN2-Cbi directly targets the catalytic domain of sGC, resembling the effect of forskolin on adenylyl cyclases. CN2-Cbi synergistically enhances sGC activation by NO-independent regulators 3-(4-amino-5-cyclopropylpyrimidine-2-yl)-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine (BAY41-2272), 4-[((4-carboxybutyl){2-[(4-phenethylbenzyl)oxy]phenethyl}amino) methyl [benzoic]-acid (cinaciguat or BAY58-2667), and 5-chloro-2-(5-chloro-thiophene-2-sulfonylamino-N-(4-(morpholine-4-sulfonyl)-phenyl)-benzamide sodium salt (ataciguat or HMR-1766). BAY41-2272 and CN2-Cbi act reciprocally by decreasing the EC50 values. CN2-Cbi increases intracellular cGMP levels and displays vasorelaxing activity in phenylephrine-constricted rat aortic rings in an endothelium-independent manner. Both effects are synergistically potentiated by BAY41-2272. These studies uncover a new mode of sGC regulation and provide a new tool for understanding the mechanism of sGC activation and function. CN2-Cbi also offers new possibilities for its therapeutic applications in augmenting the effect of other sGC-targeting drugs.
Footnotes
This work was supported by the National Institutes of Health National Heart, Lung, and Blood Institute [Grants HL088128, 3R01-HL088128]; and the American Heart Association, South Central Affiliate [Grant-in-Aid 09GRNT2060182].
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
↵ The online version of this article (available at http://jpet.aspetjournals.org) contains supplemental material.
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ABBREVIATIONS:
- NO
- nitric oxide
- sGC
- soluble guanylyl cyclase
- HNOX
- heme nitric oxide and oxygen
- BAY58-2667
- cinaciguat
- HMR1766
- ataciguat
- DEA
- 2,2-diethyl-1-nitroso-oxthydrazine
- DMSO
- dimethyl sulfoxide
- TEA
- tetraethylammonium
- BAY41-2272
- 3-(4-amino-5-cyclopropylpyrimidine-2-yl)-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine
- ODQ
- 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one
- CN2-Cbi
- dicyanocobinamide
- Cbi
- cobinamide
- YC-1
- benzylindazole
- BAY41-8543
- 2[1(2fluorobenzyl]1Hpyrazolo [3,4b] pyridin3yl]5morpholin4yl pyrimidine4,6diamine.
- Received August 12, 2011.
- Accepted December 12, 2011.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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