Nitric oxide regulation of gene transcription via soluble guanylate cyclase and type I cGMP-dependent protein kinase.
Nitric oxide (NO) regulates the expression of multiple genes but in most cases its precise mechanism of action is unclear. We used baby hamster kidney (BHK) cells, which have very low soluble guanylate cyclase and cGMP-dependent protein kinase (G-kinase) activity, and CS-54 arterial smooth muscle cells, which express these two enzymes, to study NO regulation of the human fos promoter. The NO-releasing agent Deta-NONOate (ethanamine-2,2'-(hydroxynitrosohydrazone)bis-) had no effect on a chloramphenicol acetyltransferase (CAT) reporter gene under control of the fos promoter in BHK cells transfected with an empty vector or in cells transfected with a G-kinase Ibeta expression vector. In BHK cells transfected with expression vectors for guanylate cyclase, Deta-NONOate markedly increased the intracellular cGMP concentration and caused a small (2-fold) increase in CAT activity; the increased CAT activity appeared to be from cGMP activation of cAMP-dependent protein kinase. In BHK cells co-transfected with guanylate cyclase and G-kinase expression vectors, CAT activity was increased 5-fold in the absence of Deta-NONOate and 7-fold in the presence of Deta-NONOate. Stimulation of CAT activity in the absence of Deta-NONOate appeared to be largely from endogenous NO since we found that: (i) BHK cells produced high amounts of NO; (ii) CAT activity was partially inhibited by a NO synthase inhibitor; and (iii) the inhibition by the NO synthase inhibitor was reversed by exogenous NO. In CS-54 cells, we found that NO increased fos promoter activity and that the increase was prevented by a guanylate cyclase inhibitor. In summary, we found that NO activates the fos promoter by a guanylate cyclase- and G-kinase-dependent mechanism.
Idriss
S D
SD
Department of Pediatrics, University of California, San Diego, La Jolla, California 92093-0652, USA.
Gudi
T
T
Casteel
D E
DE
Kharitonov
V G
VG
Pilz
R B
RB
Boss
G R
GR
D000818:Animals; D002478:Cells, Cultured; D006224:Cricetinae; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism*; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000378:metabolism*; D006801:Humans; D009569:Nitric Oxide / Q000502:physiology*; D009603:Nitroso Compounds / Q000378:metabolism; D011401:Promoter Regions, Genetic; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D012995:Solubility; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D009603:Nitroso Compounds; D016760:Proto-Oncogene Proteins c-fos; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D009569:Nitric Oxide; D017868:Cyclic AMP-Dependent Protein Kinases; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases; C569718:PRKG1 protein, human; D006162:Guanylate Cyclase; D006152:Cyclic GMP