Expression of soluble guanylate cyclase activity requires both enzyme subunits.
Soluble guanylate cyclase purified from rat lung exists as a heterodimer of two subunits (70 kDa and 82 kDa). Recent cloning and sequencing of both subunit entities have revealed their primary structures. Transient expression in COS-7 cells by transfection with expression vectors containing the coding regions of the 70 kDa or the 82 kDa subunit cDNA showed no guanylate cyclase activity when cells were transfected with either subunit cDNA alone. However, a marked enzymatic activity was found after transfection with both subunits that was activated by sodium nitroprusside. The combination of separately expressed guanylate cyclase subunits could not reconstitute enzymatic activity in vitro. Furthermore, cotransfection with antisense oligonucleotides against the 70 kDa subunit or the 82 kDa subunit mRNA inhibited the guanylate cyclase activity. These data indicate that both the 70 kDa and the 82 kDa subunits must be present and interactive with each other in order to see basal guanylate cyclase activity and activation with sodium nitroprusside.
Buechler
W A
WA
Pharmaceutical Discovery, Abbott Laboratories, Illinois 60064.
Nakane
M
M
Murad
F
F
D000818:Animals; D002478:Cells, Cultured; D003600:Cytosol / Q000201:enzymology; D004247:DNA / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005786:Gene Expression Regulation; D006162:Guanylate Cyclase / Q000737:chemistry / Q000235:genetics*; D008168:Lung / Q000201:enzymology; D009599:Nitroprusside / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D012995:Solubility; D014162:Transfection
D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D009599:Nitroprusside; D004247:DNA; D006162:Guanylate Cyclase