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)

Megakaryocytes are terminally differentiated cells that, in their final hours, convert their cytoplasm into long, branched proplatelets, which remodel into blood platelets. Proplatelets elongate at an average rate of 0.85 microm/min in a microtubule-dependent process. Addition of rhodamine-tubulin to permeabilized proplatelets, immunofluorescence microscopy of the microtubule plus-end marker end-binding protein 3 (EB3), and fluorescence time-lapse microscopy of EB3-green fluorescent protein (GFP)-expressing megakaryocytes reveal that microtubules, organized as bipolar arrays, continuously polymerize throughout the proplatelet. In immature megakaryocytes lacking proplatelets, microtubule plus-ends initiate and grow by centrosomal nucleation at rates of 8.9 to 12.3 microm/min. In contrast, plus-end growth rates of microtubules within proplatelets are highly variable (1.5-23.5 microm/min) and are both slower and faster than those seen in immature cells. Despite the continuous assembly of microtubules, proplatelets continue to elongate when net microtubule assembly is arrested. One alternative mechanism for force generation is microtubule sliding. Triton X-100-permeabilized proplatelets containing dynein and its regulatory complex, dynactin, but not kinesin, elongate with the addition of adenosine triphosphate (ATP) at a rate of 0.65 microm/min. Retroviral expression in megakaryocytes of dynamitin (p50), which disrupts dynactin-dynein function, inhibits proplatelet elongation. We conclude that while continuous polymerization of microtubules is necessary to support the enlarging proplatelet mass, the sliding of overlapping microtubules is a vital component of proplatelet elongation.
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PMID:Differential roles of microtubule assembly and sliding in proplatelet formation by megakaryocytes. 1611 21

Proteins in the cytoplasmic dynein pathway accumulate at the microtubule plus end, giving the appearance of comets when observed in live cells. The targeting mechanism for NUDF (LIS1/Pac1) of Aspergillus nidulans, a key component of the dynein pathway, has not been clear. Previous studies have demonstrated physical interactions of NUDF/LIS1/Pac1 with both NUDE/NUDEL/Ndl1 and CLIP-170/Bik1. Here, we have identified the A. nidulans CLIP-170 homologue, CLIPA. The clipA deletion did not cause an obvious nuclear distribution phenotype but affected cytoplasmic microtubules in an unexpected manner. Although more microtubules failed to undergo long-range growth toward the hyphal tip at 32 degrees C, those that reached the hyphal tip were less likely to undergo catastrophe. Thus, in addition to acting as a growth-promoting factor, CLIPA also promotes microtubule dynamics. In the absence of CLIPA, green fluorescent protein-labeled cytoplasmic dynein heavy chain, p150(Glued) dynactin, and NUDF were all seen as plus-end comets at 32 degrees C. However, under the same conditions, deletion of both clipA and nudE almost completely abolished NUDF comets, although nudE deletion itself did not cause a dramatic change in NUDF localization. Based on these results, we suggest that CLIPA and NUDE both recruit NUDF to the microtubule plus end. The plus-end localization of CLIPA itself seems to be regulated by different mechanisms under different physiological conditions. Although the KipA kinesin (Kip2/Tea2 homologue) did not affect plus-end localization of CLIPA at 32 degrees C, it was required for enhancing plus-end accumulation of CLIPA at an elevated temperature (42 degrees C).
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PMID:CLIP-170 homologue and NUDE play overlapping roles in NUDF localization in Aspergillus nidulans. 1646 75

Forces in the spindle that align and segregate chromosomes produce a steady poleward flux of kinetochore microtubules (MTs [kMTs]) in higher eukaryotes. In several nonmammalian systems, flux is driven by the tetrameric kinesin Eg5 (kinesin 5), which slides antiparallel MTs toward their minus ends. However, we find that the inhibition of kinesin 5 in mammalian cultured cells (PtK1) results in only minor reduction in the rate of kMT flux from approximately 0.7 to approximately 0.5 microm/min, the same rate measured in monopolar spindles that lack antiparallel MTs. These data reveal that the majority of poleward flux of kMTs in these cells is not driven by Eg5. Instead, we favor a polar "pulling-in" mechanism in which a depolymerase localized at kinetochore fiber minus ends makes a major contribution to poleward flux. One candidate, Kif2a (kinesin 13), was detected at minus ends of fluxing kinetochore fibers. Kif2a remains associated with the ends of K fibers upon disruption of the spindle by dynein/dynactin inhibition, and these K fibers flux.
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PMID:Kinesin 5-independent poleward flux of kinetochore microtubules in PtK1 cells. 1663 43

Kinesin-2 is a major microtubule-based motor in most cell types. Its in vitro motile properties have been analyzed extensively and been found to differ considerably from kinesin-1. Although recombinant kinesin-2 heterodimers exhibit processive movement, the processivity of the native kinesin-2 holoenzyme has never been evaluated. Kinesin-2 can interact with dynactin, a 'processivity factor' for cytoplasmic dynein, which may alter its motile properties. In this study, we analyze the in vitro motility of single native kinesin-2 molecules and determine the effects of dynactin on motor processivity. We find that individual native kinesin-2 molecules travel processively. Dynactin has no effect on velocity but significantly increases the run length of kinesin-2 movements. These results show that the interaction with dynactin has important functional consequences on the activity of the kinesin-2 motor.
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PMID:Dynactin enhances the processivity of kinesin-2. 1718 72

Dynactin links cytoplasmic dynein and other motors to cargo and is involved in organizing radial microtubule arrays. The largest subunit of dynactin, p150(glued), binds the dynein intermediate chain and has an N-terminal microtubule-binding domain. To examine the role of microtubule binding by p150(glued), we replaced the wild-type p150(glued) in Drosophila melanogaster S2 cells with mutant DeltaN-p150 lacking residues 1-200, which is unable to bind microtubules. Cells treated with cytochalasin D were used for analysis of cargo movement along microtubules. Strikingly, although the movement of both membranous organelles and messenger ribonucleoprotein complexes by dynein and kinesin-1 requires dynactin, the substitution of full-length p150(glued) with DeltaN-p150(glued) has no effect on the rate, processivity, or step size of transport. However, truncation of the microtubule-binding domain of p150(glued) has a dramatic effect on cell division, resulting in the generation of multipolar spindles and free microtubule-organizing centers. Thus, dynactin binding to microtubules is required for organizing spindle microtubule arrays but not cargo motility in vivo.
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PMID:Microtubule binding by dynactin is required for microtubule organization but not cargo transport. 1732 6

Transport of cellular and neuronal vesicles, organelles, and other particles along microtubules requires the molecular motor protein dynein (Mallik and Gross, 2004). Critical to dynein function is dynactin, a multiprotein complex commonly thought to be required for dynein attachment to membrane compartments (Karki and Holzbaur, 1999). Recent work also has found that mutations in dynactin can cause the human motor neuron disease amyotrophic lateral sclerosis (Puls et al., 2003). Thus, it is essential to understand the in vivo function of dynactin. To test directly and rigorously the hypothesis that dynactin is required to attach dynein to membranes, we used both a Drosophila mutant and RNA interference to generate organisms and cells lacking the critical dynactin subunit, actin-related protein 1. Contrary to expectation, we found that apparently normal amounts of dynein associate with membrane compartments in the absence of a fully assembled dynactin complex. In addition, anterograde and retrograde organelle movement in dynactin deficient axons was completely disrupted, resulting in substantial changes in vesicle kinematic properties. Although effects on retrograde transport are predicted by the proposed function of dynactin as a regulator of dynein processivity, the additional effects we observed on anterograde transport also suggest potential roles for dynactin in mediating kinesin-driven transport and in coordinating the activity of opposing motors (King and Schroer, 2000).
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PMID:Dynactin is required for coordinated bidirectional motility, but not for dynein membrane attachment. 1736 Sep 70

The identification of the Wlds gene that delays axonal degeneration in several models of neurodegenerative disease provides an interesting tool to study mechanisms of axonal loss. We showed that crossing a mouse mutant with a motoneuron disease (pmn for progressive motor neuronopathy) with mice that express the Wlds gene delayed axonal loss, increased the life span, partially rescued axonal transport deficit and prolonged the survival of the motoneuron cell bodies. To determine factors involved in the neuroprotective effect of Wlds, we combined laser capture microdissection and microarray analysis to identify genes that are differentially regulated at a pre-symptomatic age in motoneuron cell bodies in pmn/pmn,Wlds/Wlds mice as compared with pmn/pmn mice. Only 56 genes were de-regulated; none of the 'classical' genes implicated in apoptosis were de-regulated. Interestingly, a large proportion of these genes are related to axonal function and to retrograde and anterograde transport (i.e. members of the dynactin complex and kinesin family). These results were confirmed by real-time PCR, in situ hybridization and at protein level in sciatic nerves. Thus, genes related to axonal function and in particular to axonal transport may be involved at an early stage in the neuroprotective property of the Wlds gene and confirm the importance of axonal involvement in this model of motor neuron disease.
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PMID:Axonal involvement in the Wlds neuroprotective effect: analysis of pure motoneurons in a mouse model protected from motor neuron disease at a pre-symptomatic age. 1740 73

The microtubule motor cytoplasmic dynein and its activator dynactin drive vesicular transport and mitotic spindle organization. p150(Glued) is the dynactin subunit responsible for binding to dynein and microtubules. The F-box proteins constitute one of the four subunits of ubiquitin protein ligase complex called SCFs (SKP1-cullin-F-box), which governs phosphorylation-dependent ubiquitination and subsequent proteolysis. Our recent study showed that the proteolysis of mitotic kinesin CENP-E is mediated by SCF via a direct Skp1 link [D. Liu, N. Zhang, J. Du, X. Cai, M. Zhu, C. Jin, Z. Dou, C. Feng, Y. Yang, L. Liu, K. Takeyasu, W. Xie, X. Yao, Interaction of Skp1 with CENP-E at the midbody is essential for cytokinesis, Biochem. Biophys. Res. Commun. 345 (2006) 394-402]. Here we show that F-box protein FBXL5 interacts with p150(Glued) and orchestrates its turnover via ubiquitination. FBXL5 binds to p150(Glued)in vitro and in vivo. FBXL5 and p150(Glued) co-localize primarily in the cytoplasm with peri-nuclear enrichment in HeLa cells. Overexpression of FBXL5 promotes poly-ubiquitination of p150(Glued) and protein turnover of p150(Glued). Our findings provide a potential mechanism by which p150(Glued) protein function is regulated by SCFs.
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PMID:FBXL5 interacts with p150Glued and regulates its ubiquitination. 1753 94

Impaired axonal transport may promote pathogenesis in neurodegenerative disorders, such as Alzheimer's disease (AD). We previously showed that tau, amyloid precursor protein (APP), and intracellular amyloid beta-protein (Abeta) accumulate in the nerve-ending fraction of aged monkey brains, perhaps because of impaired axonal transport. In the present study, we assessed age-related changes of axonal transport motor proteins in aged monkey brains. Western blotting showed that kinesin, dynein, and dynactin (DYN) localizations dramatically changed with aging, and dynein level in nerve-ending fractions increased significantly. Coimmunoprecipitation analyses showed that DYN-dynein intermediate chain (DIC) interactions decreased, suggesting that age-related attenuation of this interaction may cause the impairment of dynein function. Moreover, RNAi-induced down-regulation of DIC in human neuroblastoma cells caused endogenous tau and APP to accumulate, and their subcellular localizations were also affected. Our findings suggest that aging attenuates DYN-DIC interaction, representing one of the risk factors for age-related impaired dynein function and even for accumulation of disease proteins.
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PMID:Aging attenuates dynactin-dynein interaction: down-regulation of dynein causes accumulation of endogenous tau and amyloid precursor protein in human neuroblastoma cells. 1762 3

Bipolarity of the meiotic spindle, required for proper chromosome segregation, is maintained throughout cell division despite rapid microtubule turnover. How this is achieved has remained mysterious, as determining the organization of individual spindle microtubules has been difficult. Here, we develop single-fluorophore speckle imaging to examine microtubule organization in the vertebrate meiotic spindle. We find that the mean length of microtubules is approximately 40% of spindle length. Long and short filaments distribute randomly throughout the spindle and those in close proximity can move in the same direction with highly heterogeneous velocities. The ratio between microtubule and spindle lengths remains unchanged as spindles elongate upon dynein-dynactin inhibition. However, maintaining this ratio depends on proper kinesin-5 function. Our data suggest that force transmission within the spindle must be understood in terms of the crosslinking dynamics of a tiled array of individual filaments, most of which do not span the distance from the pole to the metaphase plate.
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PMID:Architectural dynamics of the meiotic spindle revealed by single-fluorophore imaging. 1797 45


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