title: N-[2-bromocinnamyl(amino)ethyl]-5-isoquinolinesulphonamide (H-89) inhibits incorporation of choline into phosphatidylcholine via inhibition of choline kinase and has no effect on the phosphorylation of CTP:phosphocholine cytidylyltransferase.
abstract: We have shown previously that N-[2-bromocinnamyl(amino)-ethyl]-5-isoquinolinesulphonamide (H-89), a selective inhibitor of cyclic-AMP-dependent protein kinase (PKA), inhibits phosphatidylcholine biosynthesis in HeLa cells. In the present study, we elucidated the mechanism underlying the described inhibition. Treatment of cells with 10 microM H-89 had no effect on the phosphorylation of CTP:phosphocholine cytidylyltransferase. However, H-89 slightly affected the distribution of cytidylyltransferase between cytosol and membranes, but the cellular 1,2-diacylglycerol content was not influenced. Furthermore, pulse-chase experiments revealed that H-89 did not affect cytidylyltransferase activity. Instead, H-89 inhibited choline kinase, the enzyme catalysing the first step in the CDP-choline pathway. In the presence of 10 microM H-89, choline kinase activity was inhibited by 36 +/- 7.6% in vitro. Additionally, the phosphorylation of choline to phosphocholine was inhibited by 30 +/- 3% in cell-culture experiments. This inhibitory effect could be partly prevented by simultaneous addition of 10 microM forskolin, indicating that choline kinase is regulated in part by PKA activity.
authors.affiliation: Institut für Molekularbiologie und Biochemie, Freie Universität Berlin, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Wieprecht
authors.forename: T
authors.initials: T
authors.lastname: Wieder
authors.forename: C C
authors.initials: CC
authors.lastname: Geilen
mesh_terms: D002462:Cell Membrane / Q000378:metabolism; D002794:Choline / Q000378:metabolism*; D002797:Choline Kinase / Q000037:antagonists & inhibitors*; D019916:Choline-Phosphate Cytidylyltransferase; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004075:Diglycerides / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology*; D009713:Nucleotidyltransferases / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D013449:Sulfonamides
chemical_list: D004075:Diglycerides; D007546:Isoquinolines; D010713:Phosphatidylcholines; D013449:Sulfonamides; D005576:Colforsin; D002797:Choline Kinase; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D002794:Choline