QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online 5 August 2008
doi: 10.1242/jcs.031195

This Article Summary Full Text Full Text (PDF) Supplementary Material Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in JCS Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hayashi, S. Articles by Shingyoji, C. Search for Related Content PubMed PubMed Citation Articles by Hayashi, S. Articles by Shingyoji, C. Social Bookmarking                         What's this?

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

Department of Biological Sciences, Graduate School of Science, University of Tokyo, Hongo, Tokyo 113-0033, Japan

View larger version (42K): [in this window] [in a new window]   Fig. 1. The regulation of dynein activity in a switching model of beating flagella and the experimental design for testing the hypothesis. (A) In the sea urchin sperm flagellum, principal and reverse bends (PB and RB) are cyclically formed in the plane of beat (white), which is perpendicular to the plane of the CP (grey). (B) The formation of PB and RB is due to P-sliding (PS) and R-sliding (RS), respectively, induced by the dynein arms of the doublets 7 and 3 (or 4) (Nakano et al., 2003). (C) A model, based on our previous study (Morita and Shingyoji, 2004), illustrating the postulated sequential regulation of dynein activity (with time and along the flagellum), which induces oscillation for a half beat cycle (from top to bottom). (D) Interpretation of the previous experiment (Morita and Shingyoji, 2004). Externally applied bending induces backward sliding between the two sets of doublets in the elastase-treated axonemal fragment when ATP was applied locally and transiently by using caged-ATP with a UV flash. The testing hypothesis is that backward sliding is caused by switching of the activity of dynein from doublet 7 to doublet 3 (in the 8-3 pattern). (E) Overview of the present experiments. The effects of the direction of externally applied bending and those of the dynein attachment states on the subsequent regulation of dynein activity are analysed by using three types (1-3) of axonemes obtained from elastase-treated quiescent flagella. Finally, the switching of dynein activity by bending is tested by measuring the velocity of microtubule sliding.

View larger version (50K): [in this window] [in a new window]   Fig. 2. Effects of imposed bending on the reversal of microtubule sliding in elastase-treated quiescent flagellar axonemes severed distally to the basal P-bend. (A,B) Video images with explanatory diagrams. UV flashes induced splitting of the flagellum into two doublet bundles. When the region of overlap was bent in the P-bend direction (left panels), the subsequent UV flashes induced backward sliding of the thinner bundle (open arrow in 5) in the whole region of the bending, whereas by imposed bending in the R-bend direction (right panels), forward sliding (filled arrows in 5) was induced. (C) Relative frequency of sliding patterns induced by imposed bending. (D) Tracings showing that, by bending the distal region of the flagellum in the P-bend direction, backward sliding in the proximal region of the bending (open arrow) and splitting of the distal edge of the thinner bundle in the bent region were induced by a UV flash. (E) Tracings taken from the same axoneme as shown in B (left) after bending.

View larger version (53K): [in this window] [in a new window]   Fig. 3. Effects of imposed bending on the reversal of microtubule sliding in elastase-treated quiescent flagellar axonemes. (A-C) Video images with explanatory diagrams. UV flashes induced splitting of the flagellum into two doublet bundles at the basal P-bend (A). Bending the distal part of the region of overlap of two bundles in the P-bend direction (B) induced further forward sliding (UV2), whereas bending in the R-bend direction (C) induced backward sliding (UV3, 4) (supplementary material Movies 1 and 2). (D) Relative frequency of sliding patterns induced by imposed bending. (E) Tracings showing that the effect of the presence of the original basal P-bend on the direction of sliding. When the basal end of the flagellum was severed (Cut) after induction of the forward sliding at the basal P-bend, and the proximal region was bent to induce a new R-bend in the more distal region of the P-bend (Bending), subsequent application of ATP (UV3) induced backward sliding (open arrows) only at the proximal part of the imposed bending. However, the distal part still continued forward sliding (filled arrows).

View larger version (44K): [in this window] [in a new window]   Fig. 4. Velocity of microtubule sliding and sliding disintegration in elastase-treated flagella. (A) Movie images with tracings (top panels) showing sliding of singlet microtubules interacted with dynein arms exposed on the thinner (left panels) and the thicker (right panels) bundles of doublets obtained by splitting between the bundles in quiescent flagella. The microtubules moved away from the head after a UV flash. (B) Distribution of sliding velocities of singlet microtubules on the thinner (left) and thicker (right) doublet bundles at 1 mM caged-ATP and 10–4 M Ca2+. The velocity on the thicker bundles was significantly lower than that on the thinner bundles. (C) Average sliding velocities with their standard deviations (bars) measured in forward (FW) and backward (BW) sliding with and without imposed bending in axonemal fragments (left graph), in the quiescent flagella severed at the basal P-bend (middle graph) and in the quiescent flagella (right graph). Asterisks indicate that the differences between the two are statistically significant (Mann-Whitney U test, **P<0.01; *P<0.05).

View larger version (22K): [in this window] [in a new window]   Fig. 5. Profiles of sliding disintegration between two doublet bundles in elastase-treated quiescent flagella with (A-D) or without (E) severing at the basal P-bend. The distance of sliding of the thinner bundle towards the base was plotted against time. B and D correspond to Fig. 2A, right and left, respectively. (F) Relative frequencies of time lag at the onset of sliding with or without imposed bending in the axonemal fragments (upper three boxes) and the quiescent flagella with or without severing at the basal P-bend (lower four boxes).

View larger version (18K): [in this window] [in a new window]   Fig. 6. Relative direction of shear forces or microtubule sliding within the axoneme during bend formation. (A) The basis for curvature controlled models for flagella by Brokaw (Brokaw, 2001) with some modifications. The black arrows indicate the direction of shear forces or of sliding in the flagellum. For bends propagating from left to right, the direction of internal sliding must reverse at the points shown by the open arrows when the curvature exceeds critical curvatures. (B) Localized cyclical bending induced by iontophoretic applications of ATP to a demembranated or to a demembranated and elastase-treated sea urchin sperm flagellum (Shingyoji and Takahashi, 1995). (C) Interpretation of the backward sliding observed in the present study. (1) In the elastase-treated quiescent flagella severed at the base (Fig. 2A,B, left), P-sliding is induced by ATP application (left). Bending in the P-bend direction (middle) induces backward sliding (right) mainly in the region proximal to the bending. (2,3), In the elastase-treated quiescent axoneme (Fig. 3C,E), P-sliding occurs in the distal part of the basal P-bend (left). Bending the distal region of the flagellum in the R-bend direction (middle) induces backward sliding in the whole region of the bending (2, right) or only in the proximal region of the bending (3).

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter