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1.
P C Bridgman 《The Journal of cell biology》1999,146(5):1045-1060
To investigate the role that myosin Va plays in axonal transport of organelles, myosin Va-associated organelle movements were monitored in living neurons using microinjected fluorescently labeled antibodies to myosin Va or expression of a green fluorescent protein-myosin Va tail construct. Myosin Va-associated organelles made rapid bi-directional movements in both normal and dilute-lethal (myosin Va null) neurites. In normal neurons, depolymerization of microtubules by nocodazole slowed, but did not stop movement. In contrast, depolymerization of microtubules in dilute-lethal neurons stopped movement. Myosin Va or synaptic vesicle protein 2 (SV2), which partially colocalizes with myosin Va on organelles, did not accumulate in dilute-lethal neuronal cell bodies because of an anterograde bias associated with organelle transport. However, SV2 showed peripheral accumulations in axon regions of dilute-lethal neurons rich in tyrosinated tubulin. This suggests that myosin Va-associated organelles become stranded in regions rich in dynamic microtubule endings. Consistent with these observations, presynaptic terminals of cerebellar granule cells in dilute-lethal mice showed increased cross-sectional area, and had greater numbers of both synaptic and larger SV2 positive vesicles. Together, these results indicate that myosin Va binds to organelles that are transported in axons along microtubules. This is consistent with both actin- and microtubule-based motors being present on these organelles. Although myosin V activity is not necessary for long-range transport in axons, myosin Va activity is necessary for local movement or processing of organelles in regions, such as presynaptic terminals that lack microtubules. 相似文献
2.
Rapid movement of microtubules in axons 总被引:1,自引:0,他引:1
Cytoskeletal and cytosolic proteins are transported along axons in the slow components of axonal transport at average rates of about 0.002-0.1 microm/s. This movement is essential for axonal growth and survival, yet the mechanism is poorly understood. Many studies on slow axonal transport have focused on tubulin, the subunit protein of microtubules, but attempts to observe the movement of this protein in cultured nerve cells have been largely unsuccessful. Here, we report direct observations of the movement of microtubules in cultured nerve cells using a modified fluorescence photobleaching strategy combined with difference imaging. The movements are rapid, with average rates of 1 microm/s, but they are also infrequent and highly asynchronous. These observations indicate that microtubules are propelled along axons by fast motors. We propose that the overall rate of movement is slow because the microtubules spend only a small proportion of their time moving. The rapid, infrequent, and highly asynchronous nature of the movement may explain why the axonal transport of tubulin has eluded detection in so many other studies. 相似文献
3.
Cytoskeletal architecture and immunocytochemical localization of microtubule-associated proteins in regions of axons associated with rapid axonal transport: the beta,beta''-iminodipropionitrile-intoxicated axon as a model system 总被引:19,自引:15,他引:4
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Axons from rats treated with the neurotoxic agent beta,beta'-iminodipropionitrile (IDPN) were examined by quick-freeze, deep-etch electron microscopy. Microtubules formed bundles in the central region of the axons, whereas neurofilaments were segregated to the periphery. Most membrane-bounded organelles, presumably including those involved in rapid axonal transport, were associated with the microtubule domain. The high resolution provided by quick-freeze, deep-etch electron microscopy revealed that the microtubules were coated with an extensive network of fine strands that served both to cross-link the microtubules and to interconnect them with the membrane-bounded organelles. The strands were decorated with granular materials and were irregular in dimension. They appeared either singly or as an extensive anastomosing network in fresh axons. The microtubule-associated strands were observed in fresh, saponin-extracted, or aldehyde-fixed tissue. To explore further the identity of the microtubule-associated strands, microtubules purified from brain tissue and containing the high molecular weight microtubule-associated proteins MAP 1 and MAP 2 were examined by quick-freeze, deep-etch electron microscopy. The purified microtubules were connected by a network of strands quite similar in appearance to those observed in the IDPN axons. Control microtubule preparations consisting only of tubulin and lacking the MAPs were devoid of associated strands. To learn which of the MAPs were present in the microtubule bundles in the axon, sections of axons from IDPN-treated rats were examined by immunofluorescence microscopy using antibodies to MAP 1A, MAP 1B, MAP 2, and tubulin. Anti-MAP 2 staining was only marginally detectable in the IDPN-treated axons, consistent with earlier observations. Anti-MAP 1A and anti-MAP 1B brightly stained the IDPN-treated axons, with the staining exclusively limited to the microtubule domains. Furthermore, thin section-immunoelectron microscopy using colloidal gold-labeled second antibodies revealed that both anti-MAP 1A and anti-MAP 1B stained fuzzy filamentous structures between microtubules. In view of earlier work indicating that rapid transport is associated with the microtubule domain in the IDPN-treated axon, it now appears that MAP 1A and MAP 1B may play a role in this process. We believe that MAP 1A and MAP 1B are major components of the microtubule-associated fibrillar matrix in the axon. 相似文献
4.
Bridgman PC 《Journal of neurobiology》2004,58(2):164-174
Axonal transport in neurons has been shown to be microtubule dependent, driven by the molecular motor proteins kinesin and dynein. However, organelles undergoing fast transport can often pause or rapidly change directions without apparent dissociation from their transport tracks. Cytoskeletal polymers such as neurofilaments and microtubules have also been shown to make infrequent but rapid movements in axons indicating that their transport is likely to involve molecular motors. In addition, neurons have multiple compartments that are devoid of microtubules where transport of organelles is still seen to occur. These areas are rich in other cytoskeletal polymers such as actin filaments. Transported organelles have been shown to associate with multiple motor proteins including myosins. This suggests that nonmicrotubule-based transport may be myosin driven. In this review we will focus our attention on myosin motors known to be present in neurons and evaluate the evidence that they contribute to transport or other functions in the different compartments of the neuron. 相似文献
5.
The organization of the axonal cytoskeleton is a key determinant of the normal function of an axon, which is a long thin projection of a neuron. Under normal conditions two axonal cytoskeletal polymers, microtubules and neurofilaments, align longitudinally in axons and are interspersed in axonal cross-sections. However, in many neurotoxic and neurodegenerative disorders, microtubules and neurofilaments segregate apart from each other, with microtubules and membranous organelles clustered centrally and neurofilaments displaced to the periphery. This striking segregation precedes the abnormal and excessive neurofilament accumulation in these diseases, which in turn leads to focal axonal swellings. While neurofilament accumulation suggests an impairment of neurofilament transport along axons, the underlying mechanism of their segregation from microtubules remains poorly understood for over 30 years. To address this question, we developed a stochastic multiscale model for the cross-sectional distribution of microtubules and neurofilaments in axons. The model describes microtubules, neurofilaments and organelles as interacting particles in a 2D cross-section, and is built upon molecular processes that occur on a time scale of seconds or shorter. It incorporates the longitudinal transport of neurofilaments and organelles through this domain by allowing stochastic arrival and departure of these cargoes, and integrates the dynamic interactions of these cargoes with microtubules mediated by molecular motors. Simulations of the model demonstrate that organelles can pull nearby microtubules together, and in the absence of neurofilament transport, this mechanism gradually segregates microtubules from neurofilaments on a time scale of hours, similar to that observed in toxic neuropathies. This suggests that the microtubule-neurofilament segregation can be a consequence of the selective impairment of neurofilament transport. The model generates the experimentally testable prediction that the rate and extent of segregation will be dependent on the sizes of the moving organelles as well as the density of their traffic. 相似文献
6.
Juliana?C. Wortman Uttam?M. Shrestha Devin?M. Barry Michael?L. Garcia Steven?P. Gross Clare?C. Yu 《Biophysical journal》2014,106(4):813-823
Long-distance intracellular axonal transport is predominantly microtubule-based, and its impairment is linked to neurodegeneration. In this study, we present theoretical arguments that suggest that near the axon boundaries (walls), the effective viscosity can become large enough to impede cargo transport in small (but not large) caliber axons. Our theoretical analysis suggests that this opposition to motion increases rapidly as the cargo approaches the wall. We find that having parallel microtubules close enough together to enable a cargo to simultaneously engage motors on more than one microtubule dramatically enhances motor activity, and thus minimizes the effects of any opposition to transport. Even if microtubules are randomly placed in axons, we find that the higher density of microtubules found in small-caliber axons increases the probability of having parallel microtubules close enough that they can be used simultaneously by motors on a cargo. The boundary effect is not a factor in transport in large-caliber axons where the microtubule density is lower. 相似文献
7.
Juliana C. Wortman Uttam M. Shrestha Devin M. Barry Michael L. Garcia Steven P. Gross Clare C. Yu 《Biophysical journal》2014
Long-distance intracellular axonal transport is predominantly microtubule-based, and its impairment is linked to neurodegeneration. In this study, we present theoretical arguments that suggest that near the axon boundaries (walls), the effective viscosity can become large enough to impede cargo transport in small (but not large) caliber axons. Our theoretical analysis suggests that this opposition to motion increases rapidly as the cargo approaches the wall. We find that having parallel microtubules close enough together to enable a cargo to simultaneously engage motors on more than one microtubule dramatically enhances motor activity, and thus minimizes the effects of any opposition to transport. Even if microtubules are randomly placed in axons, we find that the higher density of microtubules found in small-caliber axons increases the probability of having parallel microtubules close enough that they can be used simultaneously by motors on a cargo. The boundary effect is not a factor in transport in large-caliber axons where the microtubule density is lower. 相似文献
8.
Mukhopadhyay R Kumar S Hoh JH 《BioEssays : news and reviews in molecular, cellular and developmental biology》2004,26(9):1017-1025
Neurofilaments and microtubules are important components of the neuronal cytoskeleton. In axons or dendrites, these filaments are aligned in parallel arrays, and separated from one another by nonrandom distances. This distinctive organization has been attributed to cross bridges formed by NF side arms or microtubule-associated proteins. We recently proposed a polymer-brush-based mechanism for regulating interactions between neurofilaments and between microtubules. In this model, the side arms of neurofilaments and the projection domains of microtubule-associated proteins are highly unstructured and exert long-range repulsive forces that are largely entropic in origin; these forces then act to organize the cytoskeleton in axons and dendrites. Here, we review the biochemical, biophysical, genetic and cell biological data for the polymer-brush and cross-bridging models. We explore how the data traditionally used to support cross bridging may be reconciled with a polymer-brush mechanism and compare the implications of recent experimental insights into axonal transport and physiology for each model. 相似文献
9.
Redistribution of proteins of fast axonal transport following administration of beta,beta'-iminodipropionitrile: a quantitative autoradiographic study 总被引:6,自引:5,他引:1
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《The Journal of cell biology》1982,95(2):672-675
Beta,beta'-iminodipropionitrile (IDPN) produces a rearrangement of axoplasmic organelles with displacement of microtubules, smooth endoplasmic reticulum, and mitochondria toward the center and of neurofilaments toward the periphery of the axon, whereas the rate of the fast component of axonal transport is unchanged. Separation of microtubules and neurofilaments makes the IDPN axons an excellent model for study of the role of these two organelles in axonal transport. The cross-sectional distribution of [3H]-labeled proteins moving with the front of the fast transport was analyzed by quantitative electron microscopic autoradiography in sciatic nerves of IDPN-treated and control rats, 6 h after injection of a 1:1 mixture of [3H]-proline and [3H]-lysine into lumbar ventral horns. In IDPN axons most of the transported [3H] proteins were located in the central region with microtubules, smooth endoplasmic reticulum and mitochondria, whereas few or none were in the periphery with neurofilaments. In control axons the [3H]-labeled proteins were uniformly distributed within the axoplasm. It is concluded that in fast axonal transport: (a) neurofilaments play no primary role; (b) the normal architecture of the axonal cytoskeleton and the normal cross-sectional distribution of transported materials are not indispensable for the maintenance of a normal rate of transport. The present findings are consistent with the models of fast transport that envision microtubules as the key organelles in providing directionality and propulsive force to the fast component of axonal transport. 相似文献
10.
Synthesis, axonal transport, and turnover of the high molecular weight microtubule-associated protein MAP 1A in mouse retinal ganglion cells: tubulin and MAP 1A display distinct transport kinetics
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Microtubule-associated proteins (MAPs) in neurons establish functional associations with microtubules, sometimes at considerable distances from their site of synthesis. In this study we identified MAP 1A in mouse retinal ganglion cells and characterized for the first time its in vivo dynamics in relation to axonally transported tubulin. A soluble 340-kD polypeptide was strongly radiolabeled in ganglion cells after intravitreal injection of [35S]methionine or [3H]proline. This polypeptide was identified as MAP 1A on the basis of its co-migration on SDS gels with MAP 1A from brain microtubules; its co-assembly with microtubules in the presence of taxol or during cycles of assembly-disassembly; and its cross-reaction with well-characterized antibodies against MAP 1A in immunoblotting and immunoprecipitation assays. Glial cells of the optic nerve synthesized considerably less MAP 1A than neurons. The axoplasmic transport of MAP 1A differed from that of tubulin. Using two separate methods, we observed that MAP 1A advanced along optic axons at a rate of 1.0-1.2 mm/d, a rate typical of the Group IV (SCb) phase of transport, while tubulin moved 0.1-0.2 mm/d, a group V (SCa) transport rate. At least 13% of the newly synthesized MAP 1A entering optic axons was incorporated uniformly along axons into stationary axonal structures. The half-residence time of stationary MAP 1A in axons (55-60 d) was 4.6 times longer than that of MAP 1A moving in Group IV, indicating that at least 44% of the total MAP 1A in axons is stationary. These results demonstrate that cytoskeletal proteins that become functionally associated with each other in axons may be delivered to these sites at different transport rates. Stable associations between axonal constituents moving at different velocities could develop when these elements leave the transport vector and incorporate into the stationary cytoskeleton. 相似文献
11.
Luminal material in microtubules of frog olfactory axons: structure and distribution 总被引:1,自引:1,他引:0
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P R Burton 《The Journal of cell biology》1984,99(2):520-528
The substructure and distribution of luminal material in microtubules of olfactory axons were studied in the bullfrog, Rana catesbeiana. By using numerous fixation methods, with and without osmium tetroxide, the luminal component was shown not to be an artifact of fixation. The material consists of globular elements 4-5 nm in diameter loosely arranged within the lumen in a discontinuous column. Counts of microtubules showing luminal material were obtained for axons in the proximal and distal ends of the olfactory nerve, and it was found that 16-18% more of the microtubules in the distal regions showed the luminal component. This raises the possibility that the material might be translocated within the microtubule lumen and tends to accumulate as it moves distally toward the axon terminal. In contrast to those of the olfactory axons, microtubules assembled in vitro from frog brain tubulin did not show luminal material. When microtubules in olfactory axons were depolymerized in situ by cold and calcium treatment and then induced to reassemble, most of those that were formed de novo showed empty lumina. Such evidence suggests that the luminal material is not an integral component of the microtubule. The hypothesis is discussed that material may be translocated within the lumina of microtubules. Furthermore, in the case of neuronal microtubules, the possibility is raised that they may serve as conduits for their own wall subunits. 相似文献
12.
Factors involved in fast axonal transport (motor proteins, microtubules, organelles, etc.) have been identified but the molecular mechanism controlling transport is unknown. We used video enhanced microscopy to directly evaluate the effect of calcium on fast axonal transport (FAxT). FAxT alterations included rapid speed decreases (within minutes) in Ca2+ free buffer and rapid speed increases (within seconds) when axons were treated with parathyroid hormone, BAY K 8644, or K+ depolarization. The speed increases were blocked by dihydropyridine Ca2+ channel antagonists. Ryanodine (20 microM), known to block calcium release from subcellular stores, caused a decrease in the rate of retrograde FAxT. Calcium ionophore A23187 (at 1 and 20 micrograms/ml) caused increases in FAxT, an effect also noted only in retrograde moving organelle traffic. Hyper- or hypo-tonic solutions produced no alterations making axoplasmic viscosity changes an unlikely explanation for the speed changes. Reproducible alteration of FAxT by manipulation of Ca2+ levels provides evidence that Ca2+ modulates fast axonal transport. Retrograde transport appears more sensitive to changes in Ca2+ and differential effects on antero- and retro-FAxT mechanisms suggest directional specificity for some of these signals which may be based upon the organelle size. Endogenous substances (e.g. PTH) that trigger axonal Ca2+ changes may rapidly modulate the rate of material delivery in axons. The results are discussed within the context of a Ca2+/calmodulin-dependent modification of the cytoskeletal matrix. 相似文献
13.
Microtubule‐binding protein doublecortin‐like kinase 1 (DCLK1) guides kinesin‐3‐mediated cargo transport to dendrites
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Joanna Lipka Lukas C Kapitein Jacek Jaworski Casper C Hoogenraad 《The EMBO journal》2016,35(3):302-318
In neurons, the polarized distribution of vesicles and other cellular materials is established through molecular motors that steer selective transport between axons and dendrites. It is currently unclear whether interactions between kinesin motors and microtubule‐binding proteins can steer polarized transport. By screening all 45 kinesin family members, we systematically addressed which kinesin motors can translocate cargo in living cells and drive polarized transport in hippocampal neurons. While the majority of kinesin motors transport cargo selectively into axons, we identified five members of the kinesin‐3 (KIF1) and kinesin‐4 (KIF21) subfamily that can also target dendrites. We found that microtubule‐binding protein doublecortin‐like kinase 1 (DCLK1) labels a subset of dendritic microtubules and is required for KIF1‐dependent dense‐core vesicles (DCVs) trafficking into dendrites and dendrite development. Our study demonstrates that microtubule‐binding proteins can provide local signals for specific kinesin motors to drive polarized cargo transport. 相似文献
14.
Summary The lumbar sympathetic ganglia and the interganglionic interconnecting nerves of untreated rats and rats treated with Colchicine (COL) or Vinblastine (VIN) were studied with the help of the Falck-Hillarp fluorescence technique and electron microscopy. Both in untreated and drug treated rats there was a good correlation between the distribution of noradrenaline (NA) specific fluorescence and granular vesicles supporting the previous view that the granular vesicles represent the main intraneuronal NA storage sites. The granular vesicles were present both in the cell bodies—mainly in the peripheral part of the cytoplasm— and in the axons of untreated rats. After local application of COL or VIN on the ganglia there was a marked increase in fluorescence intensity and number of granular vesicles in many cell bodies. Often increased number of granular vesicles were found in the neighbourhood of the Golgi apparatus, in which region only few such vesicles are found in untreated rats. In some cell bodies high numbers of granular vesicles could be found all over the cytoplasm.When applied locally to axons the mitosis inhibitors caused a marked accumulation of fluorescence and granular vesicles—and other cell organelles like mitochondria and tubules of the endoplasmic reticulum-proximal to the site of application.A prominent feature both in cell bodies and axons of drug treated rats were large bundles of neurofilaments running through the cytoplasm. In the axons these filaments were often localized to the central part of the axon and surrounded by vesicles and tubules. Microtubules, on the other hand, which are rather numerous in cell bodies and axons of untreated rats seemed to be reduced in number after COL or VIN treatment, especially in those axons in which large amounts of subcellular organelles had accumulated.The present findings are discussed with respect to intraneuronal transport of NA and possible mechanisms behind this transport. It is suggested that the accumulation of fluorescence and granular vesicles after application of mitosis inhibitors is due to an interruption of the centrifugal transport of NA granules. The increased numbers of granular vesicles in the neighbourhood of the Golgi apparatus suggest that granular vesicles are produced in this part of the cytoplasm. This does not exclude a local formation of granular vesicles in other parts of the neuron. Furthermore, the possibility is discussed that the interruption of the transport is related to the increased number of neurofilaments and a possible decrease or disarrangement of microtubules. This discussion is based on previous suggestions that microtubules are involved in intracellular transport mechanisms and on recent findings that COL and VIN bind to proteins specific for microtubules.This study has been supported by grants from the Swedish Medical Research Council (B70-14X-2887-01; B71-14X-2887-02A; B71-14P-3262-01 A; B70-14X-2207-04; B71-14X-2207-05A; K70-40P-3045-01A), from Magnus Bergwalls Foundation, from Wilhelm and Martina Lundgrens Foundation, from the Medical Faculty, University of Göteborg.For generous supply of vinblastine (Velbe®) we thank Eli Lilly Ltd.The skilful technical assistance of Mrs Kirsten Collin, Mrs Waldraut Hiort and Mr Pär-Anders Larsson is gratefully acknowledged. 相似文献
15.
The microtubule-severing proteins spastin and katanin participate differently in the formation of axonal branches 总被引:1,自引:0,他引:1
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Yu W Qiang L Solowska JM Karabay A Korulu S Baas PW 《Molecular biology of the cell》2008,19(4):1485-1498
Neurons express two different microtubule-severing proteins, namely P60-katanin and spastin. Here, we performed studies on cultured neurons to ascertain whether these two proteins participate differently in axonal branch formation. P60-katanin is more highly expressed in the neuron, but spastin is more concentrated at sites of branch formation. Overexpression of spastin dramatically enhances the formation of branches, whereas overexpression of P60-katanin does not. The excess spastin results in large numbers of short microtubules, whereas the excess P60-katanin results in short microtubules intermingled with longer microtubules. We hypothesized that these different microtubule-severing patterns may be due to the presence of molecules such as tau on the microtubules that more strongly shield them from being severed by P60-katanin than by spastin. Consistent with this hypothesis, we found that axons depleted of tau show a greater propensity to branch, and that this is true whether or not the axons are also depleted of spastin. We propose that there are two modes by which microtubule severing is orchestrated during axonal branch formation, one based on the local concentration of spastin at branch sites and the other based on local detachment from microtubules of molecules such as tau that regulate the severing properties of P60-katanin. 相似文献
16.
Effects of lidocaine on axonal morphology,microtubules, and rapid transport in rabbit vagus In vitro
Margaret R. Byers B. Raymond Fink Ross D. Kennedy Mical E. Middaugh Anita E. Hendrickson 《Developmental neurobiology》1973,4(2):125-143
Treatment with lidocaine, at exposures which completely block impulse conduction and rapid axonal transport, resulted in a sequence of morphological effects on rabbit vagus never in vitro. Low exposures (0.3% for 90 min, 0.4% for 45 min, 0.6% for 25 min) caused a reorientation and proliferation of smooth endoplasmic reticulum (SER); the number of axonal microtubules either remained normal or was increased. Intermediate exposures (0.4% for 90 min, 0.6% for 45 min) Produced a similar effect on the SER, as well as causing a 50% reduction in microtubule number and some swelling of axons and Schwann cells. The highest exposure (0.6% for 75 min) caused over 90% reduction in microtubule number, a partial loss of neurofilaments, and severe swelling of axons, Schwann call, and mitochondria. Reversibility of these effects was tested in lidocaine-treated nerves that had been washed with fresh culture medium. There was good recovery of nerve exposures. After intermediate exposures, there was only partial return of rapid transport, even though imposures, there was morphological, and no recovery occured throughout the excised vagus nerve in efferent and afferent axons as well as Schwann cells, and these events did not coincide with changes in the functional state of rapid axonal transport. 相似文献
17.
Axoplasmic organelles move on actin as well as microtubules in vitro and axons contain a large amount of actin, but little is known about the organization and distribution of actin filaments within the axon. Here we undertake to define the relationship of the microtubule bundles typically found in axons to actin filaments by applying three microscopic techniques: laser-scanning confocal microscopy of immuno-labeled squid axoplasm; electronmicroscopy of conventionally prepared thin sections; and electronmicroscopy of touch preparations-a thin layer of axoplasm transferred to a specimen grid and negatively stained. Light microscopy shows that longitudinal actin filaments are abundant and usually coincide with longitudinal microtubule bundles. Electron microscopy shows that microfilaments are interwoven with the longitudinal bundles of microtubules. These bundles maintain their integrity when neurofilaments are extracted. Some, though not all microfilaments decorate with the S1 fragment of myosin, and some also act as nucleation sites for polymerization of exogenous actin, and hence are definitively identified as actin filaments. These actin filaments range in minimum length from 0.5 to 1.5 µm with some at least as long as 3.5 µm. We conclude that the microtubule-based tracks for fast organelle transport also include actin filaments. These actin filaments are sufficiently long and abundant to be ancillary or supportive of fast transport along microtubules within bundles, or to extend transport outside of the bundle. These actin filaments could also be essential for maintaining the structural integrity of the microtubule bundles. 相似文献
18.
Rao MV Garcia ML Miyazaki Y Gotow T Yuan A Mattina S Ward CM Calcutt NA Uchiyama Y Nixon RA Cleveland DW 《The Journal of cell biology》2002,158(4):681-693
The COOH-terminal tail of mammalian neurofilament heavy subunit (NF-H), the largest neurofilament subunit, contains 44-51 lysine-serine-proline repeats that are nearly stoichiometrically phosphorylated after assembly into neurofilaments in axons. Phosphorylation of these repeats has been implicated in promotion of radial growth of axons, control of nearest neighbor distances between neurofilaments or from neurofilaments to other structural components in axons, and as a determinant of slow axonal transport. These roles have now been tested through analysis of mice in which the NF-H gene was replaced by one deleted in the NF-H tail. Loss of the NF-H tail and all of its phosphorylation sites does not affect the number of neurofilaments, alter the ratios of the three neurofilament subunits, or affect the number of microtubules in axons. Additionally, it does not reduce interfilament spacing of most neurofilaments, the speed of action potential propagation, or mature cross-sectional areas of large motor or sensory axons, although its absence slows the speed of acquisition of normal diameters. Most surprisingly, at least in optic nerve axons, loss of the NF-H tail does not affect the rate of transport of neurofilament subunits. 相似文献
19.
Polarized kinesin‐driven transport is crucial for development and maintenance of neuronal polarity. Kinesins are thought to recognize biochemical differences between axonal and dendritic microtubules in order to deliver their cargoes to the appropriate domain. To identify kinesins that mediate polarized transport, we prepared constitutively active versions of all the kinesins implicated in vesicle transport and expressed them in cultured hippocampal neurons. Seven kinesins translocated preferentially to axons and five translocated into both axons and dendrites. None translocated selectively to dendrites. Highly homologous members of the same subfamily displayed distinctly different translocation preferences and were differentially regulated during development. By expressing chimeric kinesins, we identified two microtubule‐binding elements within the motor domain that are important for selective translocation. We also discovered elements in the dimerization domain of kinesin‐2 motors that contribute to their selective translocation. These observations indicate that selective interactions between kinesin motor domains and microtubules can account for polarized transport to the axon, but not for selective dendritic transport. 相似文献
20.
The effect of in vitro exposure of bullfrog spinal nerves to 0.2 mM chlorimipramine on the density of axonal microtubules was studied in an attempt to clarify the mechanism by which chlorimipramine inhibits fast axonal transport. A 17-h exposure to chlorimipramine reduced the density of microtubules in unmyelinated axons by only 18%; this microtubular loss does not reach the upper limit of the range of microtubule reduction associated with inhibition of fast axonal transport. A 23-h exposure to chlorimipramine, which had decreased microtubular density in unmyelinated axons by 40% in a previous study, did not decrease microtubular density in myelinated axons in the present study. These results rule out microtubular destruction as the mechanism responsible for inhibition of fast orthograde axonal transport by chlorimipramine, and greatly reduce the likelihood that microtubular destruction plays a significant role in the inhibition of fast retrograde transport by chlorimipramine. 相似文献