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Philip V Bayly - One of the best experts on this subject based on the ideXlab platform.
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apparent Flexural Rigidity of single flagella depends on inter doublet shear stiffness
Biophysical Journal, 2017Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Motile cilia and flagella are whip-like subcellular organelles that bend actively to propel cells or move fluids. Normal motile functions of cilia and flagella play a critical role in many developmental and physiological processes, while ciliary dysfunction is associated with a number of ciliopathies. Efficient bending deformation of cilia and flagella depends on coordinated interactions between active forces from an array of motor proteins and passive mechanical resistance from the complex cytoskeletal structure (the axoneme). However, details of this coordination, especially axonemal mechanics, remain unclear. We investigated two major biophysical parameters, Flexural Rigidity and inter-doublet shear stiffness, of single flagella in the unicellular alga Chlamydomonas reinhardtii. Combining theoretical analysis with optical tweezers and counterbend experiments, we demonstrated that the apparent Flexural Rigidity of the axoneme depends on both the intrinsic Flexural Rigidity and the elastic inter-doublet shear stiffness. By comparing wild-type flagella with specific structural mutants, we found that the lack of nexin-dynein regulatory complexes (N-DRC) or dynein arms significantly reduces inter-doublet shear stiffness. The quantitative understanding of axonemal mechanics will ultimately lead to the development of novel diagnostic and therapeutic methods for cilia-related disorders.
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Flexural Rigidity and shear stiffness of flagella estimated from induced bends and counterbends
Biophysical Journal, 2016Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Abstract Motile cilia and flagella are whiplike cellular organelles that bend actively to propel cells or move fluid in passages such as airways, brain ventricles, and the oviduct. Efficient motile function of cilia and flagella depends on coordinated interactions between active forces from an array of motor proteins and passive mechanical resistance from the complex cytoskeletal structure (the axoneme). However, details of this coordination, including axonemal mechanics, remain unclear. We investigated two major mechanical parameters, Flexural Rigidity and interdoublet shear stiffness, of the flagellar axoneme in the unicellular alga Chlamydomonas reinhardtii. Combining experiment, theory, and finite element models, we demonstrate that the apparent Flexural Rigidity of the axoneme depends on both the intrinsic Flexural Rigidity ( EI ) and the elastic resistance to interdoublet sliding (shear stiffness, k s ). We estimated the average intrinsic Flexural Rigidity and interdoublet shear stiffness of wild-type Chlamydomonas flagella in vivo, rendered immotile by vanadate, to be EI = 840 ± 280 pN⋅ μ m 2 and k s = 79.6 ± 10.5 pN/rad, respectively. The corresponding values for the pf3; cnk11-6 double mutant, which lacks the nexin-dynein regulatory complex (N-DRC), were EI = 1011 ± 183 pN· μ m 2 and k s = 39.3 ± 6.0 pN/rad under the same conditions. Finally, in the pf13A mutant, which lacks outer dynein arms and inner dynein arm c, the estimates were EI = 777 ± 184 pN· μ m 2 and k s = 43.3 ± 7.7 pN/rad. In the two mutant strains, the Flexural Rigidity is not significantly different from wild-type ( p > 0.05), but the lack of N-DRC (in pf3; cnk11-6 ) or dynein arms (in pf13A ) significantly reduces interdoublet shear stiffness. These differences may represent the contributions of the N-DRCs (∼40 pN/rad) and residual dynein interactions (∼35 pN/rad) to interdoublet sliding resistance in these immobilized Chlamydomonas flagella.
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Flexural Rigidity and shear stiffness of flagella
Biophysical Journal, 2012Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Cilia are thin subcellular organelles that line airways and other passages and bend actively to propel fluid and foreign materials. The ciliary cytoskeleton (the axoneme) consists of nine outer microtubule doublets surrounding a central pair of singlet microtubules. Large bending deformations of the axoneme involve relative sliding of the outer doublets driven by the motor protein dyneins. The genetics and cell biology of the ciliary structure and function have been studied extensively, but the mechanics of the axoneme remain unclear. In this study, we used the unicellular alga Chlamydomonas reinhardtii as the model system for their flagellum replicates the highly conserved molecular structure of the ciliary axoneme. Piconewton forces were applied perpendicularly on the tip of a single flagellum (length L) through a microsphere trapped in optical tweezers. Dividing the force (P) by the corresponding deflection of the flagellar tip (δ) yields the Flexural stiffness of the flagellum (K = P/δ), which was then used to calculate the apparent Flexural Rigidity (EI = KL3/3). The contributions of major structural components to passive mechanical properties were quantified by testing on flagella of specific mutations. The average apparent Flexural Rigidity of wild-type, pf-3 (without nexin links), and pf-13 flagella (without outer dynein arms) was about 2700 ± 1100, 1300 ± 550, and 650 ± 140 pN·μm2, respectively. In addition, the ratio of elastic shear stiffness (resistance to interdoublet sliding) to true Flexural Rigidity was estimated by the counterbend response in bent flagella manipulated with a glass microneedle. The quantitative understanding of axonemal mechanics will help illuminate the roles of certain genes and molecular structures in the normal and abnormal axoneme.
Enrique De La Cruz - One of the best experts on this subject based on the ideXlab platform.
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Cofilin increases the bending flexibility of actin filaments: implications for severing and cell mechanics.
Journal of Molecular Biology, 2008Co-Authors: Brannon Mccullough, Laurent Blanchoin, Jean-louis Martiel, Enrique De La CruzAbstract:We determined the Flexural (bending) rigidities of actin and cofilactin filaments from a cosine correlation function analysis of their thermally driven, two-dimensional fluctuations in shape. The persistence length of actin filaments is 9.8 microm, corresponding to a Flexural Rigidity of 0.040 pN microm(2). Cofilin binding lowers the persistence length approximately 5-fold to a value of 2.2 microm and the filament Flexural Rigidity to 0.0091 pN microm(2). That cofilin-decorated filaments are more flexible than native filaments despite an increased mass indicates that cofilin binding weakens and redistributes stabilizing subunit interactions of filaments. We favor a mechanism in which the increased flexibility of cofilin-decorated filaments results from the linked dissociation of filament-stabilizing ions and reorganization of actin subdomain 2 and as a consequence promotes severing due to a mechanical asymmetry. Knowledge of the effects of cofilin on actin filament bending mechanics, together with our previous analysis of torsional stiffness, provide a quantitative measure of the mechanical changes in actin filaments associated with cofilin binding, and suggest that the overall mechanical and force-producing properties of cells can be modulated by cofilin activity.
J Howard - One of the best experts on this subject based on the ideXlab platform.
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Flexural Rigidity of microtubules and actin filaments measured from thermal fluctuations in shape
Journal of Cell Biology, 1993Co-Authors: Frederick Gittes, B. Mickey, John Nettleton, J HowardAbstract:Microtubules are long, proteinaceous filaments that perform structural functions in eukaryotic cells by defining cellular shape and serving as tracks for intracellular motor proteins. We report the first accurate measurements of the Flexural Rigidity of microtubules. By analyzing the thermally driven fluctuations in their shape, we estimated the mean Flexural Rigidity of taxol-stabilized microtubules to be 2.2 x 10(-23) Nm2 (with 6.4% uncertainty) for seven unlabeled microtubules and 2.1 x 10(-23) Nm2 (with 4.7% uncertainty) for eight rhodamine-labeled microtubules. These values are similar to earlier, less precise estimates of microtubule bending stiffness obtained by modeling flagellar motion. A similar analysis on seven rhodamine-phalloidin-labeled actin filaments gave a Flexural Rigidity of 7.3 x 10(-26) Nm2 (with 6% uncertainty), consistent with previously reported results. The Flexural Rigidity of these microtubules corresponds to a persistence length of 5,200 microns showing that a microtubule is rigid over cellular dimensions. By contrast, the persistence length of an actin filament is only approximately 17.7 microns, perhaps explaining why actin filaments within cells are usually cross-linked into bundles. The greater Flexural Rigidity of a microtubule compared to an actin filament mainly derives from the former's larger cross-section. If tubulin were homogeneous and isotropic, then the microtubule's Young's modulus would be approximately 1.2 GPa, similar to Plexiglas and rigid plastics. Microtubules are expected to be almost inextensible: the compliance of cells is due primarily to filament bending or sliding between filaments rather than the stretching of the filaments themselves.
Manfred Schliwa - One of the best experts on this subject based on the ideXlab platform.
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Flexural Rigidity of microtubules measured with the use of optical tweezers
Journal of Cell Science, 1996Co-Authors: Harald Felgner, Rainer Frank, Manfred SchliwaAbstract:The Flexural Rigidity of single microtubules is measured using optical tweezers. Two new methods are presented. In both the optical forces of the laser trap are used to directly manipulate microtubules grown off the ends of Chlamydomonas axonemes. The shapes of the microtubules are observed by video microscopy as the hydrodynamic forces of viscous flow counteract the elastic restoring forces when the microtubules are moved actively relative to the surrounding buffer medium. To determine the Flexural Rigidity, the bending of a microtubule is analyzed under a given velocity distribution along its length. Microtubules incubated with taxol after polymerization are measured to be more flexible than those without taxol added. On the other hand, MAPs are shown to increase microtubule stiffness.
Kate S Wilson - One of the best experts on this subject based on the ideXlab platform.
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apparent Flexural Rigidity of single flagella depends on inter doublet shear stiffness
Biophysical Journal, 2017Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Motile cilia and flagella are whip-like subcellular organelles that bend actively to propel cells or move fluids. Normal motile functions of cilia and flagella play a critical role in many developmental and physiological processes, while ciliary dysfunction is associated with a number of ciliopathies. Efficient bending deformation of cilia and flagella depends on coordinated interactions between active forces from an array of motor proteins and passive mechanical resistance from the complex cytoskeletal structure (the axoneme). However, details of this coordination, especially axonemal mechanics, remain unclear. We investigated two major biophysical parameters, Flexural Rigidity and inter-doublet shear stiffness, of single flagella in the unicellular alga Chlamydomonas reinhardtii. Combining theoretical analysis with optical tweezers and counterbend experiments, we demonstrated that the apparent Flexural Rigidity of the axoneme depends on both the intrinsic Flexural Rigidity and the elastic inter-doublet shear stiffness. By comparing wild-type flagella with specific structural mutants, we found that the lack of nexin-dynein regulatory complexes (N-DRC) or dynein arms significantly reduces inter-doublet shear stiffness. The quantitative understanding of axonemal mechanics will ultimately lead to the development of novel diagnostic and therapeutic methods for cilia-related disorders.
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Flexural Rigidity and shear stiffness of flagella estimated from induced bends and counterbends
Biophysical Journal, 2016Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Abstract Motile cilia and flagella are whiplike cellular organelles that bend actively to propel cells or move fluid in passages such as airways, brain ventricles, and the oviduct. Efficient motile function of cilia and flagella depends on coordinated interactions between active forces from an array of motor proteins and passive mechanical resistance from the complex cytoskeletal structure (the axoneme). However, details of this coordination, including axonemal mechanics, remain unclear. We investigated two major mechanical parameters, Flexural Rigidity and interdoublet shear stiffness, of the flagellar axoneme in the unicellular alga Chlamydomonas reinhardtii. Combining experiment, theory, and finite element models, we demonstrate that the apparent Flexural Rigidity of the axoneme depends on both the intrinsic Flexural Rigidity ( EI ) and the elastic resistance to interdoublet sliding (shear stiffness, k s ). We estimated the average intrinsic Flexural Rigidity and interdoublet shear stiffness of wild-type Chlamydomonas flagella in vivo, rendered immotile by vanadate, to be EI = 840 ± 280 pN⋅ μ m 2 and k s = 79.6 ± 10.5 pN/rad, respectively. The corresponding values for the pf3; cnk11-6 double mutant, which lacks the nexin-dynein regulatory complex (N-DRC), were EI = 1011 ± 183 pN· μ m 2 and k s = 39.3 ± 6.0 pN/rad under the same conditions. Finally, in the pf13A mutant, which lacks outer dynein arms and inner dynein arm c, the estimates were EI = 777 ± 184 pN· μ m 2 and k s = 43.3 ± 7.7 pN/rad. In the two mutant strains, the Flexural Rigidity is not significantly different from wild-type ( p > 0.05), but the lack of N-DRC (in pf3; cnk11-6 ) or dynein arms (in pf13A ) significantly reduces interdoublet shear stiffness. These differences may represent the contributions of the N-DRCs (∼40 pN/rad) and residual dynein interactions (∼35 pN/rad) to interdoublet sliding resistance in these immobilized Chlamydomonas flagella.
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Flexural Rigidity and shear stiffness of flagella
Biophysical Journal, 2012Co-Authors: Kate S Wilson, Ruth J Okamoto, Jinyu Shao, Susan K Dutcher, Philip V BaylyAbstract:Cilia are thin subcellular organelles that line airways and other passages and bend actively to propel fluid and foreign materials. The ciliary cytoskeleton (the axoneme) consists of nine outer microtubule doublets surrounding a central pair of singlet microtubules. Large bending deformations of the axoneme involve relative sliding of the outer doublets driven by the motor protein dyneins. The genetics and cell biology of the ciliary structure and function have been studied extensively, but the mechanics of the axoneme remain unclear. In this study, we used the unicellular alga Chlamydomonas reinhardtii as the model system for their flagellum replicates the highly conserved molecular structure of the ciliary axoneme. Piconewton forces were applied perpendicularly on the tip of a single flagellum (length L) through a microsphere trapped in optical tweezers. Dividing the force (P) by the corresponding deflection of the flagellar tip (δ) yields the Flexural stiffness of the flagellum (K = P/δ), which was then used to calculate the apparent Flexural Rigidity (EI = KL3/3). The contributions of major structural components to passive mechanical properties were quantified by testing on flagella of specific mutations. The average apparent Flexural Rigidity of wild-type, pf-3 (without nexin links), and pf-13 flagella (without outer dynein arms) was about 2700 ± 1100, 1300 ± 550, and 650 ± 140 pN·μm2, respectively. In addition, the ratio of elastic shear stiffness (resistance to interdoublet sliding) to true Flexural Rigidity was estimated by the counterbend response in bent flagella manipulated with a glass microneedle. The quantitative understanding of axonemal mechanics will help illuminate the roles of certain genes and molecular structures in the normal and abnormal axoneme.