Regulation of methionine adenosyltransferase catalytic activity and messenger RNA in SH-SY5Y human neuroblastoma cells.
The human neuroblastoma cell line SH-SY5Y was used to study the regulation of methionine adenosyltransferase (MAT II; E.C.2.5.1.6.) catalytic activity and transcript levels in cells of neuronal origin. The cells were exposed for 24 hr to a medium containing different concentrations of methionine (MAT substrate) as well as medium deficient of methionine. Furthermore, cells were treated with hydroxycobalamin, SAM, and the competitive MAT inhibitor cycloleucine. The MAT catalytic activity was inversely correlated to methionine concentrations, e.g. MAT Vmax increased 2-fold in cells grown in methionine-deficient medium as compared with cells cultured under standard conditions. Interestingly, MAT Km also increased from 9.04 +/- 0.44 to 12.08 +/- 0.83 in the methionine-deficient medium. Hydroxycobalamin caused an increase in activity at 40 microM while a decrease was observed at higher concentrations (100, 200, and 400 microM). Cycloleucine caused a significant inhibition of MAT catalytic activity, i.e. the inhibition was approximately 50% in the presence of 4 mM cycloleucine. The relevance of these results for the understanding of observations on MAT catalytic activity in brains of patients with Alzheimer's disease is discussed.
Gomes Trolin
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C
Department of Medical Pharmacology, Uppsala University, BMC, Sweden.
Ekblom
J
J
Oreland
L
L
D000544:Alzheimer Disease / Q000201:enzymology; D015152:Blotting, Northern; D002384:Catalysis; D003515:Cycloleucine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D006879:Hydroxocobalamin / Q000494:pharmacology; D008716:Methionine Adenosyltransferase / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D003515:Cycloleucine; D012436:S-Adenosylmethionine; D008716:Methionine Adenosyltransferase; D006879:Hydroxocobalamin