Mutants with outer dynein arm defects or deficiencies all show a major reduction in beat frequency to about half the normal value; some of these mutants show an additional decrease in sliding velocity associated with reduced shear amplitude and an additional reduction in beat frequency, as well as other more minor modifications of the normal forward mode bending pattern. New mutants (ida98, pf30), which appear to be deficient in a subset of inner dynein arms show a reduction in sliding velocity that is primarily associated with a reduction in shear amplitude, with only a small reduction in beat frequency. These differences in motility phenotype between inner and outer dynein arm mutants suggest that inner and outer dynein arms may have distinct functions. The relatively large decrease in sliding velocity associated with partial loss of inner arms is consistent with earlier observations on pf23, a nonmotile mutant lacking inner arms, suggesting that inner arms may have an essential function in motility. The ability to generate reverse mode bending patterns is retained in some inner or outer dynein arm mutants, but appears to be decreased in those mutants which show reduced shear amplitude for the forward mode bending pattern.

Asymmetrical bending waves can be obtained by reactivating demembranated sea urchin spermatozoa at high Ca" concentrations . Moving-film flash photography shows that asymmetrical flagellar bending waves are associated with premature termination of the growth of the bends in one direction (the reverse bends) while the bends in the opposite direction (the principal bends) grow for. one full beat cycle, and with unequal rates of growth of principal and reverse bends . The relative proportions of these two components of asymmetry are highly variable .The increased angle in the principal bend is compensated by a decreased angle in the reverse bend, so that there is no change in mean bend angle; the wavelength and beat frequency are also independent of the degree of asymmetry . This new information is still insufficient to identify a particular mechanism for Ca 21 -induced asymmetry .When a developing bend stops growing before initiation of growth of a new bend in the same direction, a modification of the sliding between tubules in the distal portion of the flagellum is required. This modification can be described as a superposition of synchronous sliding on the metachronous sliding associated with propagating bending waves . Synchronous sliding is particularly evident in highly asymmetrical flagella, but is probably not the cause of asymmetry . The control of metachronous sliding appears to be unaffected by the superposition of synchronous sliding .KEY WORDS calcium " flagellamicrotubules -motility " spermatozoa A variety of situations has been found in which the movements of cilia and flagella can be modified, in vitro and presumably in vivo, by free Ca 21 ion concentrations in the range of 10' to 10 -5 M . In the case of sea urchin spermatozoa, reactivated demembranated spermatozoa swim in circular paths of decreasing radius as the Ca 21 concentration is increased (12) and the flagellar beat pattern becomes highly asymmetrical . No functional significance for this behavior is known for sea urchin J . CELT, BIOLOGY

Resact, a peptide of known sequence isolated from the jelly layer of Arbacia punctulata eggs, is a potent chemoattractant for A. punctulata spermatozoa. The chemotactic response is concentration dependent, is abolished by pretreatment of the spermatozoa with resact, and shows an absolute requirement for millimolar external calcium. A. punctulata spermatozoa do not respond to speract, a peptide isolated from the jelly layer of Strongylocentrotus purpuratus eggs. This is the first report of animal sperm chemotaxis in response to a defined egg-derived molecule.

This paper investigates the accuracy of the resistive-force theory (Gray and Hancock method) which is commonly used for hydrodynamic analysis of swimming flagella. We made a comparison between the forces, bending moments, and shear moments calculated by resistive-force theory and by the more accurate slender-body theory for large-amplitude, planar wave forms computed for a flagellar model. By making an upward empirical adjustment, by about 35%, of the classical drag coefficient values used in the resistive-force theory calculations, we obtained good agreement between the distributions of the forces and moments along the length of the flagellum predicted by the two methods when the flagellum has no cell body attached. After this adjustment, we found the rate of energy expenditure calculated by the two methods for the few typical test cases to be almost identical. The resistive-force theory is thus completely satisfactory for use in analysis of mechanisms for the control of flagellar bending, at the current level of sophistication of this analysis. We also examined the effects of the presence of a cell body attached to one end of the flagellum, which modifies the flow field experienced by the flagellum. This interaction, which is not considered in resistive-force theory, is probably insignificant for small cell bodies, such as the heads of simple spermatozoa, but for larger cell bodies, or cell bodies that have large-amplitude motions transverse to the swimming direction, use of slender-body theory is required for accurate analysis.

The mutation uni-1 gives rise to uniflagellate Chlamydomonas cells which rotate around a fixed point in the microscope field, so that the flagellar bending pattern can be photographed easily . This has allowed us to make a detailed analysis of the wild-type flagellar bending pattern and the bending patterns of flagella on several mutant strains. Cells containing uni-1, and recombinants of uni-1 with the suppressor mutations, suppf-1 and sup,,-3, show the typical asymmetric bending pattern associated with forward swimming in Chlamydomonas, although suppf-1 flagella have about one-half the normal beat frequency, apparently as the result of defective function of the outer dynein arms . The pf-17 mutation has been shown to produce nonmotile flagella in which radial spoke heads and five characteristic axonemal polypeptides are missing. Recombinants containing pf-17 and either suppf -1 or suppf -3 have motile flagella, but still lack radial-spoke heads and the associated polypeptides . The flagellar bending pattern of these recombinants lacking radial-spoke heads is a nearly symmetric, large amplitude pattern which is quite unlike the wild-type pattern . However, the presence of an intact radial-spoke system is not required to convert active sliding into bending and is not required for bend initiation and bend propagation, since all of these processes are active in the suppf pf-17 recombinants . The function of the radial-spoke system appears to be to convert the symmetric bending pattern displayed by these recombinants into the asymmetric bending pattern required for efficient swimming, by inhibiting the development of reverse bends during the recovery phase of the bending cycle.Chlamydomonas has proven to be a valuable organism for genetic and biochemical investigations of flagellar structure .Flagellar function in Chlamydomonas has received more limited attention, in part because of the difficulty of observing or photographically recording the activity of flagella on swimmin g cells. We describe here an analysis of the movement of Chlamydomonas flagella, which has been facilitated by making use of a mutant uni-1 (B . Huang, Z . Ramanis, S . Dutcher, and D . J . L. Luck, manuscript in preparation) . In uni-1 a high proportion of cells are uniflagellate. When flagella beat in the asymmetric or "breast-stroke" mode which propels biflagellate cells forward, uni-1 cells rotate, and in most cases the cells show little precession. The flagellar beat is executed in a plane perpendicular to the axis of rotation, and the bending cycle of cells in which the bending plane is stabilized by proximity to the surface of a microscope slide can be recorded easily by stroboscopic dark-field microscopy. We have used this method to analyze the normal, wild-type, "breast-stroke" mode of flagellar beating, and one type of variant beating pattern associated with a flagellar motility mutation.Among the "paralyzed" flagellar mutants isolated fromChlamydomonas reinhardtii, which show little or no flagellar motility, are several muta...

Bending of cilia and flagella results from sliding between the microtubular outer doublets, driven by dynein motor enzymes. This review reminds us that many questions remain to be answered before we can understand how dynein-driven sliding causes the oscillatory bending of cilia and flagella. Does oscillation require switching between two distinct, persistent modes of dynein activity? Only one mode, an active forward mode, has been characterized, but an alternative mode, either inactive or reverse, appears to be required. Does switching between modes use information from curvature, sliding direction, or both? Is there a mechanism for reciprocal inhibition? Can a localized capability for oscillatory sliding become self-organized to produce the metachronal phase differences required for bend propagation? Are interactions between adjacent dyneins important for regulation of oscillation and bend propagation? Cell Motil. Cytoskeleton 66: 425-436, 2009. '

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ABSTACT Two-state models for muscle cross-bridges, of the type originally detailed by Andrew Huxley, were examined. Rate functions for cross-bridge attachment and detachment can be chosen which yield a steady-state force-velocity relationship appropriate for the spontaneous generation of oscillatory contractions. Computer simulations have been used to demonstrate oscillation of such cross-bridge systems, and to demonstrate that distribution of this type of local shear oscillation along the length of a flagellum is sufficient for the generation of propagated bending waves. Many biological movement rhythms are generated by controlling elements such as the nervous system, for many locomotory and respiratory rhythms in animals, or the cell membrane, for the rhythmic beating of vertebrate heart muscle. Here, I am concerned with cases where oscillation results from intrinsic properties of the molecular mechanisms for generating movement, when coupled to an appropriate load. An intrinsic origin of oscillatory contraction has been demonstrated in insect fibrillar flight muscle, by observation that the contractions are not synchronized with nerve or muscle cell membrane excitation (1) and that glycerol-extracted muscles supplied with ATP oscillate when coupled to an appropriate load (2). The oscillatory capability of insect fibrillar muscle has been ascribed to a delayed stretch-activation of actin-myosin interaction (1, 3, 4), but the molecular mechanisms for such activation have not been detailed. Evidence for similar properties has been found in several other striated muscles (5-7).An intrinsic origin of oscillatory bending and bending wave propagation has been demonstrated in flagella, by the observation that isolated, glycerol-demembranated, flagella supplied with ATP generate all the components of normal movement-bending, oscillation, and coordinated propagation of bending waves (8,9). Distribution of oscillatory capability throughout the length of a flagellum has been demonstrated by observation of localized oscillatory bending in short regions locally activated with ATP (10). A possible similarity between the oscillatory mechanisms of flagella and insect fibrillar muscle was suggested at an early stage in the investigation of fibrillar muscle (11). Machin (12, 13) explored this possibility in quantitative terms, with an analysis of wave propagation by a flagellar model in which bending was generated by stretch-activated contractile elements on either side of the flagellum. Machin's model contains a feedback loop, with active contraction causing bending of the flagellum and stretch of the antagonistic contractile elements, coupled to stretch-activation of active contraction. If stretch-activation includes a time delay, spontaneous oscillations should result.There is now firm evidence (14, 15) that flagellar bending is generated not by an active contraction mechanism, but by an active sliding process involving interaction between adjacent elements in the ring of nine microtubular doublets which is the dominant stru...