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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)

Microtubule dynamics vary during the cell cycle, and microtubules appear to be more dynamic in vivo than in vitro. Proteins that promote dynamic instability are therefore central to microtubule behavior in living cells. Here, we report that a yeast protein of the highly conserved EB1 family, Bim1p, promotes cytoplasmic microtubule dynamics specifically during G1. During G1, microtubules in cells lacking BIM1 showed reduced dynamicity due to a slower shrinkage rate, fewer rescues and catastrophes, and more time spent in an attenuated/paused state. Human EB1 was identified as an interacting partner for the adenomatous polyposis coli (APC) tumor suppressor protein. Like human EB1, Bim1p localizes to dots at the distal ends of cytoplasmic microtubules. This localization, together with data from electron microscopy and a synthetic interaction with the gene encoding the kinesin Kar3p, suggests that Bim1p acts at the microtubule plus end. Our in vivo data provide evidence of a cell cycle-specific microtubule-binding protein that promotes microtubule dynamicity.
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PMID:Yeast Bim1p promotes the G1-specific dynamics of microtubules. 1035 17

Many cell types contain a subset of long-lived, 'stable' microtubules that differ from dynamic microtubules in that they are enriched in post-translationally detyrosinated tubulin (Glu-tubulin). Elevated Glu tubulin does not stabilize the microtubules and the mechanism for the stability of Glu microtubules is not known. We used detergent-extracted cell models to investigate the nature of Glu microtubule stability. In these cell models, Glu microtubules did not incorporate exogenously added tubulin subunits on their distal ends, while >70% of the bulk microtubules did. Ca(2+)-generated fragments of Glu microtubules incorporated tubulin, showing that Glu microtubule ends are capped. Consistent with this, Glu microtubules in cell models were resistant to dilution-induced breakdown. Known microtubule end-associated proteins (EB1, APC, p150(Glued) and vinculin focal adhesions) were not localized on Glu microtubule ends. ATP, but not nonhydrolyzable analogues, induced depolymerization of Glu microtubules in cell models. Timelapse and photobleaching studies showed that ATP triggered subunit loss from the plus end. ATP breakdown of Glu microtubules was inhibited by AMP-PNP and vanadate, but not by kinase or other inhibitors. Additional experiments showed that conventional kinesin or kif3 were not involved in Glu microtubule capping. We conclude that Glu microtubules are stabilized by a plus-end cap that includes an ATPase with properties similar to kinesins.
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PMID:Detyrosinated (Glu) microtubules are stabilized by an ATP-sensitive plus-end cap. 1105 78

Kinesins are microtubule-based motor proteins that transport cargo to specific locations within the cell. However, the mechanisms by which cargoes are directed to specific cellular locations have remained elusive. Here, we investigated the in vivo movement of the Schizosaccharomyces pombe kinesin Tea2 to establish how it is targeted to microtubule tips and cell ends. Tea2 is loaded onto microtubules in the middle of the cell, in close proximity to the nucleus, and then travels using its intrinsic motor activity primarily at the tips of polymerizing microtubules. The microtubule-associated protein Mal3, an EB1 homologue, is required for loading and/or processivity of Tea2 and this function can be substituted by human EB1. In addition, the cell-end marker Tea1 is required to anchor Tea2 to cell ends. Movement of Tea1 and the CLIP170 homologue Tip1 to cell ends is abolished in Tea2 rigor (ATPase) mutants. We propose that microtubule-based transport from the vicinity of the nucleus to cell ends can be precisely regulated, with Mal3 required for loading/processivity, Tea2 for movement and Tea1 for cell-end anchoring.
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PMID:Targeted movement of cell end factors in fission yeast. 1289 67

Post-Golgi carriers of various newly synthesized axonal membrane proteins, which possess kinesin (KIF5)-driven highly processive motility, were transported from the TGN directly to axons. We found that KIF5 has a preference to the microtubules in the initial segment of axon. Low dose paclitaxel treatment caused missorting of KIF5, as well as axonal membrane proteins to the tips of dendrites. Microtubules in the initial segment of axons showed a remarkably high affinity to EB1-YFP, which was known to bind the tips of growing microtubules. These findings revealed unique features of the microtubule cytoskeletons in the initial segment, and suggested that they provide directional information for polarized axonal transport.
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PMID:Microtubules provide directional cues for polarized axonal transport through interaction with kinesin motor head. 1297 48

Flagellar axonemes assemble and continuously turn over at the flagellar tip. The supply and removal of axonemal subunits at the tip are mediated by intraflagellar transport (IFT), a motility process essential for the assembly and maintenance of all eukaryotic flagella and cilia. IFT is characterized by the movement of large protein complexes (IFT particles) from the basal bodies to the flagellar tip by kinesin-II and from the tip back to the basal bodies by cytoplasmic dynein 1b. The IFT particles consist of approximately 16 polypeptides partitioned into two complexes, A and B, and associate with axonemal precursors/turn over products. The mechanisms by which IFT motor regulation and cargo loading/unloading occur at the flagellar tip are unknown. We identified a Chlamydomonas reinhardtii ortholog of the microtubule (MT) plus end-tracking protein EB1 [4] (CrEB1) and show here that CrEB1 localizes to the tip of flagella and to the proximal part of the basal bodies. Furthermore, we found that CrEB1 is depleted from flagella of the temperature-sensitive (ts) flagellar assembly-defective (fla) mutant fla11(ts) at the restrictive temperature. This depletion of CrEB1 is accompanied by a dramatic accumulation of IFT particle polypeptides near the flagellar tip.
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PMID:The Microtubule plus end-tracking protein EB1 is localized to the flagellar tip and basal bodies in Chlamydomonas reinhardtii. 1461 22

Microtubule dynamics underlie spindle assembly, yet we do not know how the spindle environment affects these dynamics. We developed methods for measuring two key parameters of microtubule plus-end dynamic instability in Xenopus egg extract spindles. To measure plus-end polymerization rates and localize growing plus ends, we used fluorescence confocal imaging of EB1. This revealed plus-end polymerization throughout the spindle at approximately 11 microm/min, similar to astral microtubules, suggesting polymerization velocity is not regionally regulated by the spindle. The ratio of EB1 to microtubule fluorescence revealed an enrichment of polymerizing ends near the spindle middle, indicating enhanced nucleation or rescue there. We measured depolymerization rates by creating a front of synchronized depolymerization in spindles severed with microneedles. This front could be tracked by polarization and fluorescence microscopy as it advanced from each cut edge toward the associated pole. Both imaging modalities revealed rapid depolymerization ( approximately 30 microm/min) superimposed on a subset of microtubules stable to depolymerization. Larger spindle fragments contained a higher percentage of stable microtubules, which we believe were oriented with their minus ends facing the cut. Depolymerization was blocked by the potent microtubule stabilizing agent hexylene glycol, but was unaffected by alpha-MCAK antibody and AMPPNP, which block catastrophe and kinesin motility, respectively. These measurements move us closer to understanding the complete life history of a spindle microtubule.
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PMID:Microtubule plus-end dynamics in Xenopus egg extract spindles. 1476 58

The positioning of growth sites in fission yeast cells is mediated by spatially controlled microtubule dynamics brought about by tip1p, a CLIP-170-like protein, which is localized at the microtubule tips and guides them to the cell ends. The kinesin tea2p is also located at microtubule tips and affects microtubule dynamics. Here we show that tea2p interacts with tip1p and that the two proteins move with high velocity along the microtubules toward their growing tips. There, tea2p and tip1p accumulate in larger particles. Particle formation requires the EB1 homolog, mal3p. Our results suggest a model in which kinesins regulate microtubule growth by transporting regulatory factors such as tip1p to the growing microtubule tips.
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PMID:Tea2p kinesin is involved in spatial microtubule organization by transporting tip1p on microtubules. 1517 23

Regulation of microtubule polymerization and depolymerization is required for proper cell development. Here, we report that two proteins of the Drosophila melanogaster kinesin-13 family, KLP10A and KLP59C, cooperate to drive microtubule depolymerization in interphase cells. Analyses of microtubule dynamics in S2 cells depleted of these proteins indicate that both proteins stimulate depolymerization, but alter distinct parameters of dynamic instability; KLP10A stimulates catastrophe (a switch from growth to shrinkage) whereas KLP59C suppresses rescue (a switch from shrinkage to growth). Moreover, immunofluorescence and live analyses of cells expressing tagged kinesins reveal that KLP10A and KLP59C target to polymerizing and depolymerizing microtubule plus ends, respectively. Our data also suggest that KLP10A is deposited on microtubules by the plus-end tracking protein, EB1. Our findings support a model in which these two members of the kinesin-13 family divide the labour of microtubule depolymerization.
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PMID:Functionally distinct kinesin-13 family members cooperate to regulate microtubule dynamics during interphase. 1572 56

Microtubules (MTs) are dynamic polymers that undergo cell cycle and position-sensitive regulation of polymerization and depolymerization. Although many different factors that regulate MT dynamics have been described, to date there has been no systematic analysis of genes required for MT dynamics in a single system. Here, we use a transgenic EB1::GFP strain, which labels the growing plus ends of MTs, to analyze the growth rate, nucleation rate, and distribution of growing MTs in the Caenorhabditis elegans embryo. We also present the results from an RNAi screen of 40 genes previously implicated in MT-based processes. Our findings suggest that fast microtubule growth is dependent on the amount of free tubulin and the ZYG-9-TAC-1 complex. Robust MT nucleation by centrosomes requires AIR-1, SPD-2, SPD-5, and gamma-tubulin. However, we found that centrosomes do not nucleate MTs to saturation; rather, the depolymerizing kinesin-13 subfamily member KLP-7 is required to limit microtubule outgrowth from centrosomes.
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PMID:Identification and characterization of factors required for microtubule growth and nucleation in the early C. elegans embryo. 1605 29

Adenomatous polyposis coli protein (APC) translocates to, and stabilizes, the plus-ends of microtubules. In microtubule-dependent cellular protrusions, APC frequently accumulates in peripheral clusters at the basal membrane. APC targeting to membrane clusters is important for cell migration, but the localization mechanism is poorly understood. In this study, we performed deletion mapping and defined a minimal sequence (amino acids 1-2226) that efficiently targets APC to membrane clusters. This sequence lacks DLG-1 and EB1 binding sites, suggesting that these partners are not absolutely required for APC membrane targeting. A series of APC sequences were transiently expressed in cells and compared for their ability to compete endogenous APC at the membrane; potent inhibition of endogenous APC targeting was elicited by the Armadillo- (binds KAP3A, B56alpha, and ASEF) and beta-catenin-binding domains. The Armadillo domain was predicted to inhibit APC membrane localization through sequestration of the kinesin-KAP3A complex. The role of beta-catenin in APC membrane localization was unexpected but affirmed by overexpressing the APC binding sequence of beta-catenin, which similarly reduced APC membrane staining. Furthermore, we used RNA interference to show that loss of beta-catenin reduced APC at membrane clusters in migrating cells. In addition, we report that transiently expressed APC-yellow fluorescent protein co-localized with beta-catenin, KAP3A, EB1, and DLG-1 at membrane clusters, but only beta-catenin stimulated APC anchorage at the membrane. Our findings identify beta-catenin as a regulator of APC targeting to membrane clusters and link these two proteins to cell migration.
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PMID:Membrane localization of adenomatous polyposis coli protein at cellular protrusions: targeting sequences and regulation by beta-catenin. 1662 92


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