Induction of Beating by Imposed Bending or Mechanical Pulse in Demembranated, Motionless Sea Urchin Sperm Flagella at Very Low ATP Concentrations

Abstract

ABSTRACT. A basic feature of the movement of eukaryotic flagella is oscillation. Although flagellar oscillation is thought to be regulated by a self-regulatory feedback system including the mechanical signal of bending itself, the mechanism regulating the dynein motile activity to produce oscillation is not well understood. To elucidate the mechanism, we developed a new experimental system which allowed us to analyze the conditions necessary for the induction of oscillation. When a mechanical signal of bending or a pulse was applied by micromanipulation to a demembranated motionless sea urchin sperm flagellar axoneme at very low ATP concentrations (1-3 µM), a localized pair of bends was induced. The bend formation was often followed by further responses including propagation of the distal bend of paired bends, growth and propagation of the paired bends, and cyclical beating. The beating was induced at 2.0 µM or higher concentrations of ATP, but appeared even at 1.5 µM ATP if a few µM of ADP was also present. When the proximal half of a flagellum was attached to a microneedle, beating could not be induced in the distal free region at 2 µM ATP. These results suggest that mechanical signal is involved in the mechanism regulating the motile activity of dynein to produce oscillation. Our results also showed that the presence of a small amount of ADP and the axial difference along the flagellum are factors essential for the induction of flagellar oscillation.

Mechanism of flagellar oscillation–bending-induced switching of dynein activity in elastase-treated axonemes of sea urchin sperm

“…6C1,2) may constitute a basis for the coordination of cyclical bend formation and propagation. This idea is supported by the previous finding that cyclical bend formation and propagation can be induced by imposed bending so as to form a pair of opposite bends in demembranated nonmotile sperm flagella at 2-3 μM ATP (Ishikawa and Shingyoji, 2007). The influence of the distal bend to the proximal sliding is also indicated by the previous study showing that immotile bull sperm flagellum in the presence of Ni 2+ recovers its beating by bending the distal region in the direction opposite to the original basal bend (Lindemann et al, 1995).…”

“…As the regulation of flagellar motility is basically coupled with the mechanical bending itself (Hayashibe et al, 1997;Holcomb-Wygle et al, 1999;Ishikawa and Shingyoji, 2007;Morita and Shingyoji, 2004;Okuno and Hiramoto, 1976;Shingyoji et al, 1991), in order to understand the mechanism of flagellar oscillation, an analysis of the effects of bending on microtubule sliding seems indispensable. The switching hypothesis (Satir, 1985), proposed as a pioneering model to explain the regulation of dynein activity in beating flagella and cilia, postulates alternate activation of the dynein arms on either half of the axoneme beside the CP.…”

“…Flagella can respond to externally applied bending by changing the way their microtubule sliding is regulated (Eshel and Gibbons, 1989;Okuno and Hiramoto, 1976;. In addition, bending an immotile flagellum can stimulate its dynein to start beating (Hayashibe et al, 1997;Ishikawa and Shingyoji, 2007;Okuno and Hiramoto, 1976). Thus, it is likely that the flagellar bending itself is involved in the regulation of dynein activity, which would imply that a selfregulatory feedback mechanism underlies flagellar oscillation.…”

Flagellar and ciliary beating: the proven and the possible

“…By looking at individual doublet pairs sticking out from the frayed end of a Chlamydomonas flagellum, they showed that the pairs of doublets set up a limited beat of their own by sliding and bending to a critical bend angle and then separating near the base where the t-force is greatest. Recent important studies have demonstrated that imposed bending can initiate and reverse episodes of sliding in sea urchin sperm flagella when the doublets have been freed to slide by enzymatically cutting the nexin links with elastase (Morita and Shingyoji, 2004;Hayashi and Shingyoji, 2008); moreover, imposed bending can initiate beating at very low ATP concentrations (Ishikawa and Shingyoji, 2007). These results demonstrated that the curvature imposed on the partially digested structure controls the direction of dynein-driven sliding as the axoneme disintegrates.…”

Mechanism Regulating Ca2+-dependent Mechanosensory Behaviour in Sea Urchin Spermatozoa

“…In the unicellular algae Chlamydomonas reinhardtii, mechanical agitation of the cell suspension is followed by Ca 2+ -dependent activation of flagellar motility (Wakabayashi et al, 2009). However, the effects of MS on the behaviour of sperm have not been well documented, although the effects on the movement of flagella themselves have been demonstrated (Okuno and Hiramoto, 1976;Ishikawa and Shingyoji, 2007).…”