Colocalization of prostacyclin synthase with prostaglandin H synthase-1 (PGHS-1) but not phorbol ester-induced PGHS-2 in cultured endothelial cells.
The subcellular colocalization of prostacyclin synthase (PGIS) with prostaglandin H synthase (PGHS) has not been delineated. To test the hypothesis that its colocalization with PGHS is crucial for prostacyclin synthesis, we determined subcellular locations of PGIS, PGHS-1, and PGHS-2 in bovine aortic endothelial cells by immunofluorescent confocal microscopy. PGIS and PGHS-1 were colocalized to nuclear envelope (NE) and endoplasmic reticulum (ER) in resting and adenovirus-infected bovine aortic endothelial cells. PGIS and PGHS-2 were also colocalized to ER in serum-treated or adenovirus-cyclooxygenase-2-infected cells. By contrast, PGIS was not colocalized with PGHS-2 in cells induced with phorbol 12-myristate 13-acetate where PGHS-2 was visualized primarily in vesicle-like structures. The lack of colocalization was accompanied by failed prostacyclin production. Resting ECV304 cells did not produce prostacyclin and had no detectable PGHS-1 and PGIS proteins. Confocal analysis showed abnormal colocalization of PGIS and PGHS-1 to a filamentous structure. Interestingly, the abundant PGIS and PGHS-1 expressed in adenovirus-infected ECV304 cells were colocalized to NE and ER, which synthesized a large quantity of prostacyclin. These findings underscore the importance of colocalization of PGHS and PGIS to ER and NE in prostacyclin synthesis.
Liou
J Y
JY
Institute of Biomedical Sciences, Academia Sinica, 128 Academic Rd. Sec. 2, Taipei 115, Taiwan.
Shyue
S K
SK
Tsai
M J
MJ
Chung
C L
CL
Chu
K Y
KY
Wu
K K
KK
D000818:Animals; D001011:Aorta; D001095:Arachidonic Acids / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology*; D004790:Enzyme Induction / Q000187:drug effects; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000032:analysis* / Q000235:genetics; D007527:Isoenzymes / Q000032:analysis* / Q000096:biosynthesis* / Q000235:genetics; D008565:Membrane Proteins; D018613:Microscopy, Confocal; D009685:Nuclear Envelope / Q000201:enzymology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000032:analysis* / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000032:analysis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
D001095:Arachidonic Acids; D007527:Isoenzymes; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D003577:Cytochrome P-450 Enzyme System; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D019746:Intramolecular Oxidoreductases; C022461:prostacyclin synthetase; D013755:Tetradecanoylphorbol Acetate