Evidence for a role of 13S axonemal ATPase in modulation of ciliary microtubule sliding.
We recently demonstrated that elevated concentrations (greater than 20 microM) of the dynein substrate MgATP2- inhibit the spontaneous ATP-induced sliding disintegration of isolated, Triton-demembranated Tetrahymena cilia. We have used a turbidimetric assay (delta A350 nm) and electron microscopy to examine the effect of ATP on sliding disintegration when activated by other divalent cations. Mg2+, Ca2+, and Mn2+ are each capable of activating sliding, but only with Mg2+ and Mn2+ is disintegration inhibited by elevated ATP concentrations (greater than or equal to 1 mM). The two major ATPase activities obtained by KCl extraction of Tetrahymena axonemes differ in their cation specificities such that Mg2+ and Ca2+ activate the 21S dynein ATPase with equal efficiency, whereas the 13S axonemal ATPase activity is reduced by approximately 50% when CaATP2- replaces MgATP2- as substrate. With 1 mM MgATP2- as substrate, 10(-7) to 10(-2) M added CaCl2 alleviates the ATP-dependent inhibition of disintegration and likewise represses 13S MgATPase activity. In contrast, free Ca2+ has no effect on either the disintegration response or Mg-ATPase activity. In contrast to Triton-treated cilia, glycerinated cilia, which beat in 1 mM MgATP2-, are inhibited from beating by high CaATP2- concentrations. These substrate specificities suggest that concentration-dependent, substrate inhibition of sliding disintegration may be a manifestation of a physiological mechanism that is mediated by the 13S axonemal ATPase and that may function to modulate sliding during bend formation. However, the effects of added CaCl2 probably do not reflect a physiological mechanism for regulating beat parameters, but rather may result from CaATP2- competing for MgATP2- binding sites on the 13S ATPase, thereby blocking expression of the 13S ATPase.
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002465:Cell Movement; D002923:Cilia / Q000201:enzymology*; D004305:Dose-Response Relationship, Drug; D008274:Magnesium / Q000378:metabolism; D008870:Microtubules / Q000502:physiology*; D008970:Molecular Weight; D013379:Substrate Specificity; D013768:Tetrahymena
D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D008274:Magnesium; D002118:Calcium