The disruption of brain microtubules in vitro by the phospholipase inhibitor p-bromophenacyl bromide.
The influence of p-bromophenacyl bromide (pBPAB) and structural analogues on the assembly and Ca2+ sensitivity of porcine brain microtubules (MTs) was studied by spectrophotometric measurements in vitro. MT assembly was inhibited by 36 microM pBPAB but not by the structural analogues p-chlorophenacyl chloride or acetophenone. In the presence of pBPAB, but not structural analogues, the addition of 10 mM Ca2+ induced aggregation of polymerized MT protein, whereas a decrease in turbidity (due to MT disassembly) was observed in controls. The effects of pBPAB on both MT assembly and Ca2+ sensitivity were blocked by glutathione, but not by N-acetyl L-cysteine, N-acetyl L-lysine nor L-tyrosine, indicating that a highly reduced sulphydryl group(s) may be involved. Western blotting analyses of drug-treated MTs revealed a form of tubulin with altered electrophoretic characteristics, probably caused by a covalent interaction with pBPAB. MT preparations polymerized in the presence of the drug contained fewer MTs than control samples, the predominant structures being identified as amorphous aggregates of MT proteins. The fact that pBPAB affects MT integrity at an effective anti-inflammatory dose in vitro may reflect the involvement of MT disruption in some of the pharmacological effects of this drug. pBPAB is not therefore a suitable tool for studying the specific involvement of phospholipase A2 in cellular events.
Hargreaves
A J
AJ
Department of Pharmacology and Therapeutics, University of Liverpool, U.K.
Glazier
A P
AP
Flaskos
J
J
Mullins
F H
FH
McLean
W G
WG
D000098:Acetophenones / Q000494:pharmacology*; D000818:Animals; D001921:Brain / Q000187:drug effects* / Q000648:ultrastructure; D002118:Calcium / Q000494:pharmacology; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000737:chemistry; D008870:Microtubules / Q000187:drug effects* / Q000648:ultrastructure; D010741:Phospholipases A / Q000037:antagonists & inhibitors*; D054467:Phospholipases A2; D013552:Swine; D014404:Tubulin / Q000737:chemistry
D000098:Acetophenones; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D010741:Phospholipases A; D054467:Phospholipases A2; C006821:4-bromophenacyl bromide; C038699:acetophenone; D002118:Calcium