The Counterbend Phenomenon in Dynein-Disabled Rat Sperm Flagella and What It Reveals about the Interdoublet Elasticity

Abstract

Rat sperm that have been rendered passive by disabling the dynein motors with 50 muM sodium metavanadate and 0.1 mM ATP exhibit an interesting response to imposed bending. When the proximal flagellum is bent with a microprobe, the portion of the flagellum distal to the probe contact point develops a bend in the direction opposite the imposed bend. This "counterbend" is not compatible with a simple elastic beam. It can be satisfactorily explained by the sliding tubule model of flagellar structure but only if there are permanent elastic connections between the outer doublets of the axoneme. The elastic component that contributes the bending torque for the counterbend does not reset to a new equilibrium position after an imposed bend but returns the flagellum to a nearly straight or slightly curved final position after release from the probe. This suggests it is based on fixed, rather than mobile, attachments. It is also disrupted by elastase or trypsin digestion, confirming that it is dependent on a protein linkage. Adopting the assumption that the elasticity is attributed to the nexin links that repeat at 96 nm intervals, we find an apparent elasticity for each link that ranges from 1.6 to 10 x 10(-5) N/m. However, the elasticity is nonlinear and does not follow Hooke's law but appears to decrease with increased stretch. In addition, the responsible elastic elements must be able to stretch to more than 10 times their resting length without breakage to account for the observed counterbend formation. Elasticity created by some type of protein unfolding may be the only viable explanation consistent with both the extreme capacity for extension and the nonlinear character of the restoring force that is observed.

The geometric clutch at 20: stripping gears or gaining traction?

“…Mechanical studies of flagellar stiffness seemed to support this view. When flagella are in the relaxed condition, which is induced by either vanadate inhibition of the dynein (Lindemann et al 2005, Pelle et al 2009) or inhibition of dynein by 10 mM ATP (Lindemann et al 1973), the flagella are in their most flexible state and their stiffness is minimized.…”

Thinking about flagellar oscillation

“…not understood, but it probably includes both the effects of dynein attachment to substrate sites, and interdoublet linkages, often referred to as nexin linkages. Nexin linkages also provide an elastic resistance to sliding, which can regulate bending [Brokaw, 1980;Lindemann et al, 2005] and may be part of the mechanism for oscillation. The other interaction is at the basal end of the axoneme, where there are structural modifications that introduce a strong resistance to sliding.…”

“…In simple cilia and flagella such as those that propel sea urchin spermatozoa, there is no measurable sliding between outer doublets at the basal end. The situation in mammalian sperm flagella is more complicated [see Lindemann et al, 2005]. Outer doublets are flexible, but appear to have a high resistance to compression and extension, so the most important consequence of the basal sliding resistance is that sliding between outer doublets necessarily results in bending of the axoneme [Satir, 1968].…”

Functional anatomy of the mammalian sperm flagellum

“…If a flagellum in this condition is bent with a microprobe, the portion of the flagellum beyond the probe exhibits a bend in the opposite direction of the imposed bend; a phenomenon we termed a counterbend (Lindemann et al, 2005). The counterbend results from the fact that the outer doublets are linked to each other by elastic resistances, known as the dynein regulatory complex (drc)/nexin links (Warner, 1976;Heuser et al, 2009), that act to restore the flagellum to the straight position when shear is present.Both sea urchin sperm and mammalian sperm show the counterbend phenomenon (Lindemann et al, 2005;Pelle et al, 2009). In a sea urchin flagellum, the amount of distal shear in the counterbend, as measured by the shear angle of the tip of the flagellum, never exceeds the shear imposed on the proximal flagellum.…”