The Journal of Cell Biology, Vol 118, 1177-1188, Copyright © 1992 by The Rockefeller University Press

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The motile beta/IC1 subunit of sea urchin sperm outer arm dynein does not form a rigor bond

AG Moss, JL Gatti and GB Witman
Cell Biology Group and Male Fertility Program, Worcester Foundation for Experimental Biology, Shrewsbury, Massachusetts 01545.

We used in vitro translocation and cosedimentation assays to study the microtubule binding properties of sea urchin sperm outer arm dynein and its beta/IC1 subunit. Microtubules glided on glass-absorbed sea urchin dynein for a period of time directly proportional to the initial MgATP2- concentration and then detached when 70-95% of the MgATP2- was hydrolyzed. Detachment resulted from MgATP2- depletion, because (a) perfusion with fresh buffer containing MgATP2- reconstituted binding and gliding, (b) microtubules glided many minutes with an ATP- regenerating system at ATP concentrations which alone supported gliding for only 1-2 min, and (c) microtubules detached upon total hydrolysis of ATP by an ATP-removal system. The products of ATP hydrolysis antagonized binding and gliding; as little as a threefold excess of ADP/Pi over ATP resulted in complete loss of microtubule binding and translocation by the beta/IC1 subunit. In contrast to the situation with sea urchin dynein, microtubules ceased gliding but remained bound to glass-absorbed Tetrahymena outer arm dynein when MgATP2- was exhausted. Cosedimentation assays showed that Tetrahymena outer arm dynein sedimented with microtubules in an ATP-sensitive manner, as previously reported (Porter, M.E., and K. A. Johnson. J. Biol. Chem. 258: 6575-6581). However, the beta/IC1 subunit of sea urchin dynein did not cosediment with microtubules in the absence of ATP. Thus, this subunit, while capable of generating motility, lacks both structural and rigor-type microtubule binding.
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