Enzyme activity and protein of multiple forms of choline acetyltransferase: effects of calyculin A and okadaic acid.
Choline acetyltransferase (ChAT) appears to exist in multiple forms, three of which can be isolated biochemically as cytosolic (cChAT), ionically-membrane bound (ibChAT) and non-ionic membranous (mChAT). In this study, we first examined whether the quantitative distribution of enzyme protein and enzyme activity was the same. Enzyme activity and ChAT protein distributed similarly: the majority of ChAT activity and protein were found in cChAT followed by mChAT and least activity and amount were in ibChAT. Our second objective was to investigate the effects of calyculin A or okadaic acid on the subcellular distribution of ChAT activity and amount from rat hippocampal formation. Calyculin A and okadaic acid decreased significantly (p < 0.01) cytosolic and membranous ChAT activity; ionically-bound ChAT was not significantly (p > 0.67) different from control. Removal of calyculin A or okadaic acid restored cytosolic ChAT activity (p > 0.9 as compared to control), but not membranous enzyme activity (p < 0.05 as compared to control). The immunoreactive cytosolic ChAT was reduced significantly (p < 0.01) by calyculin A and okadaic acid. Enzyme amount of membranous ChAT was decreased significantly by calyculin A (p < 0.01) and okadaic acid (p < 0.001). Enzyme amount of ionically-bound ChAT was not changed (p > 0.99) by either of these two phosphatase inhibitors. This investigation demonstrates that alterations in ChAT activity of each subfraction parallel changes in enzyme amounts in the same fractions.
Issa
A M
AM
Department of Pharmacology and Therapeutics, McGill University, Montréal, Québec, Canada.
Gauthier
S
S
Collier
B
B
D000818:Animals; D002795:Choline O-Acetyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006624:Hippocampus / Q000187:drug effects / Q000201:enzymology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008297:Male; D008387:Marine Toxins; D019319:Okadaic Acid / Q000494:pharmacology*; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000187:drug effects / Q000201:enzymology
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D008387:Marine Toxins; D010080:Oxazoles; D019319:Okadaic Acid; C059041:calyculin A; D002795:Choline O-Acetyltransferase; D010749:Phosphoprotein Phosphatases