Analysis of Ni(2+)-induced arrest of Paramecium axonemes.
This study examines the molecular basis for paralysis of ciliary motility by Ni2+. At concentrations above 0.1 mM, Ni2+ slowed and subsequently stopped swimming of living, axenically grown Paramecium tetraurelia. However, some cilia still beat in the presence of 0.1 mM Ni2+. When permeabilized and reactivated with 4 mM ATP at pCa greater than 7, cells resumed ciliary beat and swam forward at approximately 170 +/- 28 microns s-1; swimming speed increased in the presence of 10 microM cyclic AMP. Addition of Ni2+ (pNi less than 5) caused rapid arrest of all ciliary beat in a single position. This was fully reversible when EGTA was added to raise the pNi. Axonemes were then isolated and sliding was observed in the presence of trypsin and ATP. When pNi was lowered to about 5, sliding was reduced dramatically. This too was reversible with EGTA. Dynein was then extracted from the axonemes and used for in vitro translocation assays. At concentrations of Ni2+ where microtubule-sliding and axonemal beat were greatly inhibited or absent, microtubule translocation in vitro by 22 S dynein was only slightly affected. However, translocation by 14 S dynein was stopped completely. When pNi was raised by repeated washing with solutions containing EGTA, microtubule translocation by 14 S dynein resumed. We conclude that Ni2+ induces a reversible paralysis by a direct effect on 14 S dynein while 22 S dynein is not a primary target.
Larsen
J
J
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
Satir
P
P
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D002413:Cations, Divalent / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D002923:Cilia / Q000187:drug effects; D004398:Dyneins / Q000032:analysis; D004533:Egtazic Acid; D008855:Microscopy, Electron, Scanning; D008870:Microtubules / Q000187:drug effects; D009532:Nickel / Q000494:pharmacology*; D016805:Paramecium tetraurelia / Q000737:chemistry / Q000187:drug effects*
D002413:Cations, Divalent; D004533:Egtazic Acid; D009532:Nickel; D000251:Adenosine Triphosphatases; D004398:Dyneins