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)

Ncd is a kinesin-related microtubule motor protein required for chromosome segregation in Drosophila oocytes and early embryos. In tests for interactions with other proteins, we find that mutants of alpha Tub67C, which affect an oocyte- and early embryo-specific alpha-tubulin, enhance meiotic nondisjunction and zygotic loss of ncdD, a partial loss-of-function mutant of ncd. The enhancement is dominant and allele-specific with respect to alpha Tub67C, and depends on the recessive effects of ncdD. Cytologically, embryos of alpha Tub67C/+ show delayed meiotic divisions and defective female pronucleus formation, while meiotic spindle assembly is abnormal in embryos of ncdD/ncdD. Doubly mutant alpha Tub67C ncdD/ncdD embryos are rescued for female pronucleus formation, but show delayed meiotic progression and defective pronuclear conjugation or fusion. Delayed completion of meiosis, together with failure of pronuclear fusion, prevents normal interactions of maternal with paternal chromosomes, enhancing the ncdD mutant phenotype. The genetics and cytology of doubly mutant embryos and the molecular defect of NcdD provide evidence for interaction of Ncd with alpha Tub67C in vivo. These results imply that a specific alpha-tubulin isoform is required for normal cellular function of a kinesin motor protein.
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PMID:Enhancement of the ncdD microtubule motor mutant by mutants of alpha Tub67C. 904 53

The kinesin motor protein family members move along microtubules with characteristic polarity. Chimeric motors containing the stalk and neck of the minus-end-directed motor, Ncd, fused to the motor domain of plus-end-directed kinesin were analyzed. The Ncd stalk and neck reversed kinesin motor polarity, but mutation of the Ncd neck reverted the chimeric motor to plus-end movement. Thus, residues or regions contributing to motor polarity must be present in both the Ncd neck and the kinesin motor core. The neck-motor junction was critical for Ncd minus-end movement; attachment of the neck to the stalk may also play a role.
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PMID:Determinants of kinesin motor polarity. 971 86

Proteins of the kinesin superfamily utilize a conserved catalytic motor domain to generate movements in a wide variety of cellular processes. In this review, we discuss the rapid expansion in our understanding of how eukaryotic cells take advantage of these proteins to generate force and movement in diverse functional contexts. We summarize several recent examples revealing that the simplest view of a kinesin motor protein binding to and translocating a cargo along a microtubule track is inadequate. In fact, this paradigm captures only a small subset of the many ways in which cells harness force production of the generation of intracellular movements and functions. We also highlight several situations where the catalytic kinesin motor domain may not be used to generate movement, but instead may be used in other biochemical and functional contexts. Finally, we review some recent ideas about kinesin motor regulation, redundancy, and cargo attachment strategies.
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PMID:The road less traveled: emerging principles of kinesin motor utilization. 1061 60

Slow axonal transport depends on an active mechanism that conveys cytosolic proteins. To investigate its molecular mechanism, we now constructed an in vitro experimental system for observation of tubulin transport, using squid giant axons. After injecting fluorescence-labeled tubulin into the axons, we monitored the movement of fluorescence by confocal laser scanning microscopy and fluorescence correlation spectroscopy. Here, from the pharmacological experiments and the functional blocking of kinesin motor protein by anti-kinesin antibody, we show that the directional movement of fluorescent profile was dependent on kinesin motor function. The fluorescent correlation function and estimated translational diffusion time revealed that tubulin molecule was transported in a unique form of large transporting complex distinct from those of stable polymers or other cytosolic protein.
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PMID:Oligomeric tubulin in large transporting complex is transported via kinesin in squid giant axons. 1105 54

The budding yeast shmoo tip is a model system for analyzing mechanisms coupling force production to microtubule plus-end polymerization/depolymerization. Dynamic plus ends of astral microtubules interact with the shmoo tip in mating yeast cells, positioning nuclei for karyogamy. We have used live-cell imaging of GFP fusions to identify proteins that couple dynamic microtubule plus ends to the shmoo tip. We find that Kar3p, a minus end-directed kinesin motor protein, is required, whereas the other cytoplasmic motors, dynein and the kinesins Kip2p and Kip3p, are not. In the absence of Kar3p, attached microtubule plus ends released from the shmoo tip when they switched to depolymerization. Furthermore, microtubules in cells expressing kar3-1, a mutant that results in rigor binding to microtubules [2], were stabilized specifically at shmoo tips. Imaging of Kar3p-GFP during mating revealed that fluorescence at the shmoo tip increased during periods of microtubule depolymerization. These data are the first to localize the activity of a minus end-directed kinesin at the plus ends of microtubules. We propose a model in which Kar3p couples depolymerizing microtubule plus ends to the cell cortex and the Bim1p-Kar9p protein complex maintains attachment during microtubule polymerization. In support of this model, analysis of Bim1p-GFP at the shmoo tip results in a localization pattern complementary to that of Kar3p-GFP.
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PMID:The minus end-directed motor Kar3 is required for coupling dynamic microtubule plus ends to the cortical shmoo tip in budding yeast. 1293 27

In order to investigate the chromosomal genotoxicity of nitrobenzene and benzonitrile, we studied the induction of micronuclei (MN) by these test compounds in V79 cells, as well as effects on the formation and stability of microtubules and on motor protein functions. No cytotoxicity was seen in V79 cell cultures in terms of Neutral red uptake after 18 h treatment with up to 1 mM nitrobenzene or 1 mM benzonitrile. Subsequently, a concentration range up to 100 micro M was used in the experiments on induction of MN. Both test compounds exhibit a weak, but definitely positive test result compared to the solvent (DMSO) control. Minimal effect concentrations of nitrobenzene and benzonitrile appeared as low as 0.01 micro M, and no-effect-concentrations were between 0.001 and 0.005 micro M. Clearly enhanced MN rates were found at 0.1 micro M and higher. Both, nitrobenzene and benzonitrile, induced mostly kinetochor (CREST)-positive micronuclei, thus characterising the chromosomal effects as aneugenic. In cell-free assays, a slight effect on tubulin assembly was observed at 1 mM nitrobenzene without addition of DMSO. Higher concentrations (5 mM) led to secondary effects. In presence of 1% DMSO, nitrobenzene exerted no detectable effect on tubulin assembly up to the solubility limit in water of about 15 mM. For benzonitrile in presence of DMSO, a clear dose-response of inhibition of tubulin assembly at 37 degrees C was seen above the no-effect-concentration of 2 mM, with an IC(50) of 13 mM and protein denaturation starting above a level of about 20 mM. The nature of the effects of nitrobenzene and benzonitrile on the association of tubulin to form microtubules was confirmed by electron microscopy. Treatment by either 5 mM nitrobenzene or 13 mM benzonitrile plus 1% DMSO left the microtubular structure intact whereas 5 mM nitrobenzene, in absence of DMSO, led to irregular cluster formations. The experiments demonstrate that both nitrobenzene and benzonitrile, in millimolar concentration ranges, may lead to interference with tubulin assembly in a cell-free system. The functionality of the tubulin-kinesin motor protein system was assessed using the microtubule gliding assay. Nitrobenzene affected the gliding velocity in a concentration-dependent manner, starting at about 7.5 micro M and reaching complete inhibition of motility at 30 micro M, whereas benzonitrile up to 200 micro M did not affect the kinesin-driven gliding velocity. The micronucleus assay data demonstrate a chromosomal endpoint of genotoxicity of nitrobenzene and benzonitrile. Aneugenic effects of both compounds occur at remarkably low concentrations, with lowest-effect-concentrations being 0.1 micro M. This points to the relevance of interactions with the cellular spindle apparatus.
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PMID:Chromosomal genotoxicity of nitrobenzene and benzonitrile. 1451 6

Male germ cell differentiation requires a highly cell-specific gene expression programme that is achieved by unique chromatin remodelling, transcriptional control, and the expression of testis-specific genes or isoforms. The regulatory processes governing gene expression in spermatogenesis have fundamentally unique requirements, including meiosis, ongoing cellular differentiation and a peculiar chromatin organization. The signalling cascades and the downstream effectors contributing to the programme of spermatogenesis are currently being unravelled, revealing the unique features of germ cell regulatory circuits. This paper reports on the unique role that CREM exerts as a master regulator. Targeted inactivation of the genes encoding CREM and ACT has been achieved. ACT selectively associates with KIF17b, a kinesin motor protein highly expressed in germ cells. It has been found that KIF17b directly determines the intracellular localization of ACT. Thus, the activity of a transcriptional co-activator is intimately coupled to the function of a kinesin via tight regulation of its intracellular localization. The conservation of these elements and of their regulatory functions in human spermatogenesis indicates that they are likely to provide important insights into understanding the molecular mechanisms of human infertility.
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PMID:Genetic control of spermiogenesis: insights from the CREM gene and implications for human infertility. 1570 96

The serotonin system and NMDA receptors (NMDARs) in prefrontal cortex (PFC) are both critically involved in the regulation of cognition and emotion under normal and pathological conditions; however, the interactions between them are essentially unknown. Here we show that serotonin, by activating 5-HT(1A) receptors, inhibited NMDA receptor-mediated ionic and synaptic currents in PFC pyramidal neurons, and the NR2B subunit-containing NMDA receptor is the primary target of 5-HT(1A) receptors. This effect of 5-HT(1A) receptors was blocked by agents that interfere with microtubule assembly, as well as by cellular knock-down of the kinesin motor protein KIF17 (kinesin superfamily member 17), which transports NR2B-containing vesicles along microtubule in neuronal dendrites. Inhibition of either CaMKII (calcium/calmodulin-dependent kinase II) or MEK/ERK (mitogen-activated protein kinase kinase/extracellular signal-regulated kinase) abolished the 5-HT(1A) modulation of NMDAR currents. Biochemical evidence also indicates that 5-HT(1A) activation reduced microtubule stability, which was abolished by CaMKII or MEK inhibitors. Moreover, immunocytochemical studies show that 5-HT(1A) activation decreased the number of surface NR2B subunits on dendrites, which was prevented by the microtubule stabilizer. Together, these results suggest that serotonin suppresses NMDAR function through a mechanism dependent on microtubule/kinesin-based dendritic transport of NMDA receptors that is regulated by CaMKII and ERK signaling pathways. The 5-HT(1A)-NMDAR interaction provides a potential mechanism underlying the role of serotonin in controlling emotional and cognitive processes subserved by PFC.
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PMID:Serotonin 5-HT1A receptors regulate NMDA receptor channels through a microtubule-dependent mechanism. 1594 77

N-Methyl-D-aspartate (NMDA) receptors (NMDARs), which play a key role in synaptic plasticity, are dynamically regulated by many signaling molecules and scaffolding proteins. Although actin cytoskeleton has been implicated in regulating NMDAR stability in synaptic membrane, the role of microtubules in regulating NMDAR trafficking and function is largely unclear. Here we show that microtubule-depolymerizing agents inhibited NMDA receptor-mediated ionic and synaptic currents in cortical pyramidal neurons. This effect was Ca(2+)-independent, required GTP, and was more prominent in the presence of high NMDA concentrations. The NR2B subunit-containing NMDA receptor was the primary target of microtubules. The effect of microtubule depolymerizers on NMDAR currents was blocked by cellular knockdown of the kinesin motor protein KIF17, which transports NR2B-containing vesicles along microtubule in neuronal dendrites. Neuromodulators that can stabilize microtubules, such as brain-derived neurotrophic factor, significantly attenuated the microtubule depolymerizer-induced reduction of NMDAR currents. Moreover, immunocytochemical studies show that microtubule depolymerizers decreased the number of surface NR2B subunits on dendrites, which was prevented by the microtubule stabilizer. Taken together, these results suggest that interfering with microtubule assembly suppresses NMDAR function through a mechanism dependent on kinesin-based dendritic transport of NMDA receptors.
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PMID:Microtubule regulation of N-methyl-D-aspartate receptor channels in neurons. 1597 19

The dimeric structure of kinesin superfamily proteins plays an important role in their motile functions and characteristics. In this study, the coiled-coil-forming property of the stalk region (192-346) of Drosophila ncd, a C-terminal kinesin motor protein, was investigated by synthesizing various peptide fragments. The alpha helicity of a set of 46-residue peptides spanning the stalk region appeared too low to form a coiled-coil dimer, probably because of insufficient continuity of the hydrophobic residues at (a and d) core positions in amphipathic heptad repeats. On the other hand, several peptides with leucine residues introduced at core positions or with extensional sequences with high alpha helicity had an advantage in coiled-coil formation. When we analyzed the thermal and urea-induced unfolding of these dimeric peptides, we identified four domains having a relatively high potential to form coiled coils. Among them, three domains on the C-terminal side of the stalk region, i.e., (252-272), (276-330), and (336-346), were in the same heptad frame, although these potential coiled-coil domains were not self-sustaining individually. This is in sharp contrast to the fragment of human kinesin, (332-369), which has an extremely high tendency toward coiled-coil formation. One of the possible triggers for coiled-coil formation of the ncd stalk region may be the interaction between the motor domain and the C-terminal part of the stalk as previously revealed by X-ray crystallography. The residues, S331 and R335, seem to act as a breaking point for alpha-helix continuity. This would make the region (336-346), as the head-stalk joint, more flexible such as seen with a plus-end-directed kinesin, if this region had no interaction with the motor domain. These characteristic differences between ncd and kinesin suggest that the nonlocally sustained coiled coil of ncd is one of the factors important for minus-end-directed motility.
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PMID:Coiled coil in the stalk region of ncd motor protein is nonlocally sustained. 1651 26


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