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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 2 (1982), S. 599-614 
    ISSN: 0886-1544
    Keywords: monoclonal antibodies to tubulin ; radioimmune assay ; immunoautoradiography ; Western blots ; immunofluorescence ; tubulin heterogeneity ; eukaryotic flagellar motility ; immunomotility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two monoclonal antibodies reactive for α-tubulin but not for β-tubulin have been prepared, characterized in terms of their relative binding to tubulins from differnt sources by a solid-phase binding assay, immunoautoradiography, and indirect immunofluorescence, and utilized to study flagellar motility. Our results demonstrate that α-tubulins from different species, and even from different tissues of the same species, are nonidentical. Especially interesting was the observation that one of the antibodies, Ab2, immunofluorescently stained microtubules of chick embryo fibroblast cells, but was completely unreactive for microtubules of rat kangaroo (PtK2) fibroblasts; a different antibody, Ab1, stained both cell types. Results of these and additional experiments clearly show that Ab1 and Ab2 recognize discrete and different epitopes on α-tubulin.Monoclonal antitubulins Ab1 and Ab2 each inhibited the bend amplitude of reactivated sea urchin spermatozoa without affecting beat frequencies or the ability of the outer doublet microtubules to slide past each other in elastase-digested models. These results, together with those obtained previously using rabbit polyclonal antitubulin antibodies [Asai and Brokaw, 1980], demonstrate that inhibition of bend amplitude is a common property of antitubulin antibodies and is not due to the binding of antibodies to one specific site on the axoneme. Our results suggest that tubulin subunit conformational changes may occur on the outer doublet lattice and may be integrally involved in the mechanism and control of flagellar bending.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 28 (1994), S. 199-204 
    ISSN: 0886-1544
    Keywords: axoneme ; cilia ; flagella ; microtubule ; motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Observations that were interpreted to provide evidence for equivalent functions of all axonemal dyneins should be reinterpreted, and models based on this assumption should be abandoned. In the future, attempts to understand the mechanisms for flagellar bending, oscillation, and bend propagation should start from the assumption that each type of axonemal dynein may have a specific function. At least three distinct functions can now be identified: bend initiation, maintenance of the angle of propagating bends, and generation of power to overcome viscous resistances. Only the last of these three functions is an outer arm dynein function. © 1994 Wiley-Liss, Inc.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 27 (1994), S. 150-160 
    ISSN: 0886-1544
    Keywords: axoneme ; bend propagation ; computer simulation ; flagella ; microtubule sliding ; motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The distinct damped, or attenuated, bending pattern observed when demembranated sperm flagella of the tunicate, Ciona, are reactivated in the presence of 2 mM Li+ has been analysed in detail. In these patterns, bends are initiated at the base of the flagellum, but die out after they start to propagate along the flagellum, so that little or no bending is seen in the distal half of the flagellum. A quantitative descriptive analysis shows that the distinctive feature of this attenuation of bending wave amplitude is an asymmetric interbend decay, or slippage, occuring, on average, only at the transitions between a reverse bend and the preceding principal bend. This attenuation is combined with a significant amount of synchronous sliding in the distal half of the flagellum and a decrease in propagation velocity of transitions between bends in the mid-region of the flagellum.Computer simulations demonstrate that the synchronous sliding in the distal half of these flagella can be an entirely passive consequence of the mechanical interaction between active sliding and bending in the basal third of the flagellum and viscous resistances to movement of the distal region of the flagellum through the fluid environment. The current computer models do not contain a mechanism for asymmetric interbend decay that can reproduce these attenuated bending patterns. © 1994 Wiley-Liss, Inc.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 32 (1995), S. 65-79 
    ISSN: 0886-1544
    Keywords: Ciona ; flagella ; motility ; tyrosine kinase ; cAMP-dependent kinase ; spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A specific peptide inhibitor of the cyclic AMP (cAMP)-dependent protein kinase (PKI-peptide) is a very effective inhibitor of the cAMP-dependent activation of motility of Ciona spermatozoa, when the PKI-peptide is present at the beginning of incubation of demembranated spermatozoa with cAMP and ATP. Under conditions where approximately 120 sec is required for full activation of motility, the window of sensitivity to the PKI-peptide lasts for only 25-30 sec. Examination of sperm pellet proteins labeled with 32P ATP during activation reveals a major 25 kDa phosphoprotein and 2 minor phosphoproteins whose phosphorylation is highly sensitive to inhibition by the PKI-peptide and essentially complete during this early phase. These sperm proteins appear to be immediate substrates for cAMP-dependent protein kinase, and phosphorylation of one or more of these appears to be required, but not sufficient, for activation of motility. The phosphorylation of other proteins is reduced or eliminated when PKI-peptide is present at the beginning of incubation, but is unaffected by later addition of PKI-peptide. Some of these substrates appear to be likely candidates for axonemal proteins that must be phosphorylated during the later stages of incubation in order to complete the activation process. This selection is based upon a high degree of inhibition by inclusion of PKI-peptide or other inhibitors at the start of the incubation process, on near-completion of their phosphorylation by the end of the 2 min incubation period required for activation of motility, and evidence that these proteins are phosphorylated during in vivo activation of motility. Although these observations suggest the presence of a second kinase activity that is upregulated by the initial activation of the cAMP-dependent protein kinase, assays using exogenous substrates have not yet been able to identify such a kinase activity. © 1995 Wiley-Liss, Inc.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 7 (1987), S. 160-168 
    ISSN: 0886-1544
    Keywords: flagella ; asymmetric beating ; sea urchin ; Ciona ; Chlamydomonas ; switching mechanism ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Time-averaged data covering six to ten beat cycles for ATP-reactivated spermatozoa of a sea urchin and Ciona, and from a uniflagellate mutant of Chlamydomonas, were analyzed to obtain parameters of oscillation and mean shear angle at each point along the flagellum. The mean shear angles usually show a sharp change near the base of Ihc flagellum. This sharp basal change in angle is correlated with perceived asymmetry in the development times of principal and reverse bends when these bends are measured directly from the asymmetric bending patterns, without subtracting out the mean shear angle. The asymmetry in development times was previously considered to be evidence against a “biased baseline” mechanism for asymmetric bending waves, in which completely symmetric bending waves develop and propagate on a curved flagellum. Our analysis now shows that the asymmetry in development times can be fully explained by the presence of a sharp static bend near the base of the flagellum, which can confuse the determination of the times of initiation of new bends at the base of the flagellum. Our reinterpretation of these data removes previous objections to the “biased baseline” mechanism for the regulation of bending wave asymmetry by calcium, and supports other evidence favoring a biased baseline mechanism, rather than a “biased switching” mechanism.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 26 (1993), S. 144-162 
    ISSN: 0886-1544
    Keywords: axoneme ; bend propagation ; flagella ; microtubule sliding ; motility ; vanadate ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Microtubule sliding associated with the bending of reactivated flagella of demembranated spermatozoa of the tunicate, Ciona, has been analyzed using a descriptive model that permits quantitation of metachronous and synchronous components of sliding. Reduced-amplitude bending waves, obtained by addition of increased salt (K acetate), lithium, or vanadate to the reactivation solutions, have been examined. Increased K acetate can decrease bend angle by as much as 70% with little change in frequency.In all cases, a decrease in the amplitude, or bend angle, of propagated bends is measured as a decrease in the metachronous component of sliding and is associated with a reduction in the growth of new bends after they begin to propagate during the second half-cycle of bend development. At higher K acetate concentrations, bend growth during the second half-cycle of bend development is very strongly reduced and may even become negative. A disparity between the rates of bend growth in the first and second half-cycles of bend development corresponds to a large amount of synchronous sliding in the distal portion of the flagellum. When the synchronous sliding component is large, the sliding velocity in a propagating bend decreases to near-0 values and may even reverse its direction as the bend propagates through the mid-region of the flagellum. Since these large perturbations of sliding velocity do not interfere with regular propagation of bends with nearly constant bend angle, the bend propagation mechanism cannot operate by metachronous control of the velocity of sliding, and is unlikely to operate by local monitoring of either the amount or velocity of sliding. These observations therefore argue against models in which active sliding is regulated by shear or sliding velocity, and make curvature-controlled models relatively more attractive.In many cases, a reduction in sliding during bend initiation (the first half-cycle of development of new bends) also contributes to the decreased amplitude of propagated bends. These changes in bend initiation are similar in both full-length flagella and in flagella shortened by breakage. The amount of sliding that occurs during bend initiation is relatively independent of the distribution of sliding between metachronous and synchronous components in the distal part of the flagellum. These observations therefore provide additional evidence that bend initiation and bend propagation are independent and separable processes. © 1993 Wiley-Liss, Inc.
    Additional Material: 16 Ill.
    Type of Medium: Electronic Resource
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