Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.4.4 (kinesin)
5,033 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mechanism(s) by which microtubule plus-end tracking proteins are targeted is unknown. In the filamentous fungus Aspergillus nidulans, both cytoplasmic dynein and NUDF, the homolog of the LIS1 protein, localize to microtubule plus ends as comet-like structures. Herein, we show that NUDM, the p150 subunit of dynactin, also forms dynamic comet-like structures at microtubule plus ends. By examining proteins tagged with green fluorescent protein in different loss-of-function mutants, we demonstrate that dynactin and cytoplasmic dynein require each other for microtubule plus-end accumulation, and the presence of cytoplasmic dynein is also important for NUDF's plus-end accumulation. Interestingly, deletion of NUDF increases the overall accumulation of dynein and dynactin at plus ends, suggesting that NUDF may facilitate minus-end-directed dynein movement. Finally, we demonstrate that a conventional kinesin, KINA, is required for the microtubule plus-end accumulation of cytoplasmic dynein and dynactin, but not of NUDF.
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PMID:Accumulation of cytoplasmic dynein and dynactin at microtubule plus ends in Aspergillus nidulans is kinesin dependent. 1268 3

Chlamydiae are pathogenic obligate intracellular bacteria with a biphasic developmental cycle that involves cell types adapted for extracellular survival (elementary bodies, EBs) and intracellular multiplication (reticulate bodies, RBs). The intracellular development of chlamydiae occurs entirely within a membrane-bound vacuole termed an inclusion. Within 2 hours after entry into host cells, Chlamydia trachomatis EBs are trafficked to the perinuclear region of the host cell and remain in close proximity to the Golgi apparatus, where they begin to fuse with a subset of host vesicles containing sphingomyelin. Here, we provide evidence that chlamydial migration from the cell periphery to the peri-Golgi region resembles host cell vesicular trafficking. Chlamydiae move towards the minus end of microtubules and aggregate at the microtubule-organizing center (MTOC). In mammalian cells the most important minus-end-directed microtubule motor is cytoplasmic dynein. Microinjection of antibodies to a subunit of cytoplasmic dynein inhibited movement of chlamydiae to the MTOC, whereas microinjection of antibodies to the plus-directed microtubule motor, kinesin, had no effect. Surprisingly, overexpression of the protein p50 dynamitin, a subunit of the dynactin complex that links vesicular cargo to the dynein motor in minus directed vesicle trafficking, did not abrogate chlamydial migration even though host vesicle transport was inhibited. Nascent chlamydial inclusions did, however, colocalize with the p150(Glued) dynactin subunit, which suggests that p150(Glued) may be required for dynein activation or processivity but that the cargo-binding activity of dynactin, supplied by p50 dynamitin subunits and possibly other subunits, is not. Because chlamydial transcription and translation were required for this intracellular trafficking, chlamydial proteins modifying the cytoplasmic face of the inclusion membrane are probable candidates for proteins fulfilling this function.
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PMID:Chlamydia trachomatis uses host cell dynein to traffic to the microtubule-organizing center in a p50 dynamitin-independent process. 1290 5

The XMAP215/Dis1 MAP family is thought to regulate microtubule plus-end assembly in part by antagonizing the catastrophe-promoting function of kin I kinesins, yet XMAP215/Dis1 proteins localize to centrosomes. We probed the mitotic function of TOGp (human homolog of XMAP215/Dis1) using siRNA. Cells lacking TOGp assembled multipolar spindles, confirming results of Gergely et al. (2003. Genes Dev. 17, 336-341). Eg5 motor activity was necessary to maintain the multipolar morphology. Depletion of TOGp decreased microtubule length and density in the spindle by approximately 20%. Depletion of MCAK, a kin I kinesin, increased MT lengths and density by approximately 20%, but did not disrupt spindle morphology. Mitotic cells lacking both TOGp and MCAK formed bipolar and monopolar spindles, indicating that TOGp and MCAK contribute to spindle bipolarity, without major effects on MT stability. TOGp localized to centrosomes in the absence of MTs and depletion of TOGp resulted in centrosome fragmentation. TOGp depletion also disrupted MT minus-end focus at the spindle poles, detected by localizations of NuMA and the p150 component of dynactin. The major functions of TOGp during mitosis are to focus MT minus ends at spindle poles, maintain centrosome integrity, and contribute to spindle bipolarity.
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PMID:TOGp, the human homolog of XMAP215/Dis1, is required for centrosome integrity, spindle pole organization, and bipolar spindle assembly. 1471 66

The bi-directional movement of pigment granules in frog melanophores involves the microtubule-based motors cytoplasmic dynein, which is responsible for aggregation, and kinesin II and myosin V, which are required for dispersion of pigment. It was recently shown that dynactin acts as a link between dynein and kinesin II and melanosomes, but it is not fully understood how this is regulated and if more proteins are involved. Here, we suggest that spectrin, which is known to be associated with Golgi vesicles as well as synaptic vesicles in a number of cells, is of importance for melanosome movements in Xenopus laevis melanophores. Large amounts of spectrin were found on melanosomes isolated from both aggregated and dispersed melanophores. Spectrin and two components of the oligomeric dynactin complex, p150(glued) and Arp1/centractin, co-localized with melanosomes during aggregation and dispersion, and the proteins were found to interact as determined by co-immunoprecipitation. Spectrin has been suggested as an important link between cargoes and motor proteins in other cell types, and our new data indicate that spectrin has a role in the specialized melanosome transport processes in frog melanophores, in addition to a more general vesicle transport.
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PMID:A role for spectrin in dynactin-dependent melanosome transport in Xenopus laevis melanophores. 1514 76

Recent research on kinesin motors has outlined the diversity of the superfamily and defined specific cargoes moved by kinesin family (KIF) members. Owing to the difficulty of purifying large amounts of native motors, much of this work has relied on recombinant proteins expressed in vitro. This approach does not allow ready determination of the complement of kinesin motors present in a given tissue, the relative amounts of different motors, or comparison of their native activities. To address these questions, we isolated nucleotide-dependent, microtubule-binding proteins from 13-day chick embryo brain. Proteins were enriched by microtubule affinity purification, then subjected to velocity sedimentation to separate the 20S dynein/dynactin pool from a slower sedimenting KIF containing pool. Analysis of the latter pool by anion exchange chromatography revealed three KIF species: kinesin I (KIF5), kinesin II (KIF3), and KIF1C (Unc104/KIF1). The most abundant species, kinesin I, exhibited the expected long range microtubule gliding activity. By contrast, KIF1C did not move microtubules. Kinesin II, the second most abundant KIF, could be fractionated into two pools, one containing predominantly A/B isoforms and the other containing A/C isoforms. The two motor species had similar activities, powering microtubule gliding at slower speeds and over shorter distances than kinesin I.
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PMID:Fractionation and characterization of kinesin II species in vertebrate brain. 1518 Aug 27

Our previous studies demonstrated that fluorescent early endocytic vesicles prepared from rat liver after injection of Texas red asialoorosomucoid contain asialoglycoprotein and its receptor and move and undergo fission along microtubules using kinesin I and KIFC2, with Rab4 regulating KIFC2 activity (J. Cell Sci. 116, 2749, 2003). In the current study, procedures to prepare fluorescent late endocytic vesicles were devised. In addition, flow cytometry was utilized to prepare highly purified fluorescent endocytic vesicles, permitting validation of microscopy-based experiments as well as direct biochemical analysis. These studies revealed that late vesicles bound to and moved along microtubules, but in contrast to early vesicles, did not undergo fission. As compared with early vesicles, late vesicles had reduced association with receptor, Rab4, and kinesin I but were highly associated with dynein, Rab7, dynactin, and KIF3A. Dynein and KIF3A antibodies inhibited late vesicle motility, whereas kinesin I and KIFC2 antibodies had no effect. Dynamitin antibodies prevented the association of late vesicles with microtubules. These results indicate that acquisition and exchange of specific motor and regulatory proteins characterizes and may regulate the transition of early to late endocytic vesicles. Flow cytometric purification should ultimately facilitate detailed proteomic analysis and mapping of endocytic vesicle-associated proteins.
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PMID:Microtubule-dependent movement of late endocytic vesicles in vitro: requirements for Dynein and Kinesin. 1518 Nov 54

Cytoplasmic dynein is a minus-end directed microtubule motor and plays important roles in the transport of various intracellular cargoes. Cytoplasmic dynein comprises two identical heavy chains and forms a dimer (double-headed dynein); the total molecular weight of the cytoplasmic dynein complex is about 1.5 million. The dynein motor domain is structurally very different from those of kinesin and myosin, and our understanding of the mechanisms of dynein energy transduction is limited mainly because of the difficulty in obtaining a sufficient quantity of purified and active cytoplasmic dynein. We purified cytoplasmic dynein, which was free from dynactin and other dynein-associated proteins. The purified cytoplasmic dynein was active in an in vitro motility assay. The controlled dialysis of the purified dynein against 4 M urea resulted in its complete dissociation into monomeric species (single-headed dynein). The separation of the dynein heads by the treatment was reversible. The MgATPase activities of the single-headed and reconstituted double-headed dynein were comparable to that of intact dynein. The double-headed dynein bundled microtubules in the absence of ATP; the single-headed dynein did not. The single-headed dynein produced in vitro microtubule-gliding motility at velocities very similar to those of double-headed dynein at various ATP concentrations. These results indicate that a single cytoplasmic dynein heavy chain is sufficient to produce robust microtubule motility. Application of the double- and single-headed dynein molecules in various assay systems will elucidate the mechanism of action of the cytoplasmic dynein.
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PMID:Dissociation of double-headed cytoplasmic dynein into single-headed species and its motile properties. 1523 58

During cell division metaphase spindles maintain constant length, whereas spindle microtubules continuously flux polewards, requiring addition of tubulin subunits at microtubule plus-ends, polewards translocation of the microtubule lattice, and removal of tubulin subunits from microtubule minus-ends near spindle poles. How these processes are coordinated is unknown. Here, we show that dynein/dynactin, a multi-subunit microtubule minus-end-directed motor complex, and NuMA, a microtubule cross-linker, regulate spindle length. Fluorescent speckle microscopy reveals that dynactin or NuMA inhibition suppresses microtubule disassembly at spindle poles without affecting polewards microtubule sliding. The observed uncoupling of these two components of flux indicates that microtubule depolymerization is not required for the microtubule transport associated with polewards flux. Inhibition of Kif2a, a KinI kinesin known to depolymerize microtubules in vitro, results in increased spindle microtubule length. We find that dynein/dynactin contribute to the targeting of Kif2a to spindle poles, suggesting a model in which dynein/dynactin regulate spindle length and coordinate flux by maintaining microtubule depolymerizing activities at spindle poles.
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PMID:Dynein/dynactin regulate metaphase spindle length by targeting depolymerizing activities to spindle poles. 1531 63

Melatonin (5-methoxy N-acetyltryptamine) is a hormone synthesized and released from the pineal gland at night, which acts on specific high affinity G-protein coupled receptors to regulate various aspects of physiology and behaviour, including circadian and seasonal responses, and some retinal, cardiovascular and immunological functions. In amphibians, such as Xenopus laevis, another role of melatonin is in the control of skin coloration through an action on melanin-containing pigment granules (melanosomes) in melanophores. In these cells, very low concentrations of melatonin activate the Mel(1c) receptor subtype triggering movement of granules toward the cell centre thus lightening skin colour. Mel(1c) receptor activation reduces intracellular cAMP via a pertussis toxin-sensitive inhibitory G-protein (Gi), but how this and other intracellular signals regulate pigment movement is not yet fully understood. However, melanophores have proven an excellent model for the study of the molecular mechanisms which coordinate intracellular transport. Melanosome transport is reversible and involves both actin- (myosin V) and microtubule-dependent (kinesin II and dynein) motors. Melanosomes retain both kinesin and dynein during anterograde and retrograde transport, but the myosin V motor seems to be recruited to melanosomes during dispersion, where it assists kinesin II in dominating dynein thus driving net dispersion. Recent work suggests an important role for dynactin in coordinating the activity of the opposing microtubule motors. The melanophore pigment aggregation response has also played a vital role in the ongoing effort to devise specific melatonin receptor antagonists. Much of what has been learnt about the parts of the melatonin molecule required for receptor binding and activation has come from detailed structure-activity data using novel melatonin ligands. Work aiming to devise ligands specific for the distinct melatonin receptor subtypes stands poised to deliver selective agonists and antagonists which will be valuable tools in understanding the role of this enigmatic hormone in health and disease.
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PMID:Melatonin, melatonin receptors and melanophores: a moving story. 1535 31

Neurons transmit long-range biochemical signals between cell bodies and distant axonal sites or termini. To test the hypothesis that signaling molecules are hitchhikers on axonal vesicles, we focused on the c-Jun NH2-terminal kinase (JNK) scaffolding protein Sunday Driver (syd), which has been proposed to link the molecular motor protein kinesin-1 to axonal vesicles. We found that syd and JNK3 are present on vesicular structures in axons, are transported in both the anterograde and retrograde axonal transport pathways, and interact with kinesin-I and the dynactin complex. Nerve injury induces local activation of JNK, primarily within axons, and activated JNK and syd are then transported primarily retrogradely. In axons, syd and activated JNK colocalize with p150Glued, a subunit of the dynactin complex, and with dynein. Finally, we found that injury induces an enhanced interaction between syd and dynactin. Thus, a mobile axonal JNK-syd complex may generate a transport-dependent axonal damage surveillance system.
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PMID:Sunday Driver links axonal transport to damage signaling. 1573 68


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