The protein connector enhancer of kinase suppressor of Ras 2(CNKSR2),present in both the postsynaptic density and cytoplasm of neurons,is a scaffolding protein with several protein-binding domains.Variants of the CNKS...The protein connector enhancer of kinase suppressor of Ras 2(CNKSR2),present in both the postsynaptic density and cytoplasm of neurons,is a scaffolding protein with several protein-binding domains.Variants of the CNKSR2 gene have been implicated in neurodevelopmental disorders,particularly intellectual disability,although the precise mechanism involved has not yet been fully understood.Research has demonstrated that CNKSR2 plays a role in facilitating the localization of postsynaptic density protein complexes to the membrane,thereby influencing synaptic signaling and the morphogenesis of dendritic spines.However,the function of CNKSR2 in the cytoplasm remains to be elucidated.In this study,we used immunoprecipitation and high-resolution liquid chromatography-mass spectrometry to identify the interactors of CNKSR2.Through a combination of bioinformatic analysis and cytological experiments,we found that the CNKSR2 interactors were significantly enriched in the proteome of the centrosome.We also showed that CNKSR2 interacted with the microtubule protein DYNC1H1 and with the centrosome marker CEP290.Subsequent colocalization analysis confirmed the centrosomal localization of CNKSR2.When we downregulated CNKSR2 expression in mouse neuroblastoma cells(Neuro 2A),we observed significant changes in the expression of numerous centrosomal genes.This manipulation also affected centrosome-related functions,including cell size and shape,cell proliferation,and motility.Furthermore,we found that CNKSR2 interactors were highly enriched in de novo variants associated with intellectual disability and autism spectrum disorder.Our findings establish a connection between CNKSR2 and the centrosome,and offer new insights into the underlying mechanisms of neurodevelopmental disorders.展开更多
TAU is a microtubule-associated protein that promotes microtubule assembly and stability in the axon.TAU is missorted and aggregated in an array of diseases known as tauopathies.Microtubules are essential for neuronal...TAU is a microtubule-associated protein that promotes microtubule assembly and stability in the axon.TAU is missorted and aggregated in an array of diseases known as tauopathies.Microtubules are essential for neuronal function and regulated via a complex set of post-translational modifications,changes of which affect microtubule stability and dynamics,microtubule interaction with other proteins and cellular structures,and mediate recruitment of microtubule-severing enzymes.As impairment of microtubule dynamics causes neuronal dysfunction,we hypothesize cognitive impairment in human disease to be impacted by impairment of microtubule dynamics.We therefore aimed to study the effects of a disease-causing mutation of TAU(P301L)on the levels and localization of microtubule post-translational modifications indicative of microtubule stability and dynamics,to assess whether P301L-TAU causes stability-changing modifications to microtubules.To investigate TAU localization,phosphorylation,and effects on tubulin post-translational modifications,we expressed wild-type or P301L-TAU in human MAPT-KO induced pluripotent stem cell-derived neurons(i Neurons)and studied TAU in neurons in the hippocampus of mice transgenic for human P301L-TAU(p R5 mice).Human neurons expressing the longest TAU isoform(2N4R)with the P301L mutation showed increased TAU phosphorylation at the AT8,but not the p-Ser-262 epitope,and increased polyglutamylation and acetylation of microtubules compared with endogenous TAU-expressing neurons.P301L-TAU showed pronounced somatodendritic presence,but also successful axonal enrichment and a similar axodendritic distribution comparable to exogenously expressed 2N4R-wildtype-TAU.P301L-TAU-expressing hippocampal neurons in transgenic mice showed prominent missorting and tauopathy-typical AT8-phosphorylation of TAU and increased polyglutamylation,but reduced acetylation,of microtubules compared with non-transgenic littermates.In sum,P301L-TAU results in changes in microtubule PTMs,suggestive of impairment of microtubule stability.This is accompanied by missorting and aggregation of TAU in mice but not in i Neurons.Microtubule PTMs/impairment may be of key importance in tauopathies.展开更多
Post-translational modification of spastin enables precise spatiotemporal control of its microtubule severing activity.However,the detailed mechanism by which spastin turnover is regulated in the context of neurite ou...Post-translational modification of spastin enables precise spatiotemporal control of its microtubule severing activity.However,the detailed mechanism by which spastin turnover is regulated in the context of neurite outgrowth remains unknown.Here,we found that spastin interacted with ubiquitin and was significantly degraded by K48-mediated poly-ubiquitination.Cullin3 facilitated spastin degradation and ubiquitination.RING-box protein 1,but not RING-box protein 2,acted synergistically with Cullin3 protein to regulate spastin degradation.Overexpression of Culin3 or BRX1 markedly suppressed spastin expression,and inhibited spastin-mediated microtubule severing and promotion of neurite outgrowth.Moreover,USP14 interacted directly with spastin to mediate its deubiquitination.USP14 overexpression significantly increased spastin expression and suppressed its ubiquitination and degradation.Although co-expression of spastin and USP14 did not enhance microtubule severing,it did increase neurite length in hippocampal neurons.Taken together,these findings elucidate the intricate regulatory mechanisms of spastin turnover,highlighting the roles of the Cullin-3–Ring E3 ubiquitin ligase complex and USP14 in orchestrating its ubiquitination and degradation.The dynamic interplay between these factors governs spastin stability and function,ultimately influencing microtubule dynamics and neuronal morphology.These insights shed light on potential therapeutic targets for neurodegenerative disorders associated with spastin defects.展开更多
Microtubules are one kind of cytoskeleton that is ubiquitous in eukaryotic cells and is essential for various biological processes, such as intracellular transport, maintenance of cell morphology and cell division. Mi...Microtubules are one kind of cytoskeleton that is ubiquitous in eukaryotic cells and is essential for various biological processes, such as intracellular transport, maintenance of cell morphology and cell division. Microtubules are dynamic structures and the basic unit of microtubules is the heterodimer composed of α-tubulin and β-tubulin. The biological function of microtubules is based on their dynamic polymerization and depolymerization. However, the nucleotide-dependent depolymerization mechanism of microtubules is still unclear. The dynamic instability of microtubules is determined by the interactions between tubulins. In this work, the interactions between tubulins in the microtubule lattice(GDP at the interdimer interface) and in the special GTP-cap structure(GTP at the interdimer interface) are systematically investigated using all-atom molecular dynamics simulation method. The analysis of the tubulin–tubulin and nucleotide–tubulin interactions and binding free energy at different interfaces of microtubule shows that the hydrolysis of GTP can affect the longitudinal interaction between α-tubulin and β-tubulin at the interdimer interface and the lateral interaction between β-tubulins. In particular, the displacement of M loop of β-tubulin induced by GTP hydrolysis weakens the lateral interaction betweenβ-tubulins. Based on these results, a nucleotide-dependent depolymerization mechanism of microtubule induced by GTP hydrolysis is proposed.展开更多
Colorectal cancer(CRC)has remained the second and the third leading cause of cancer-related death worldwide and in the United States,respectively.Although significant improvement in overall survival has been achieved,...Colorectal cancer(CRC)has remained the second and the third leading cause of cancer-related death worldwide and in the United States,respectively.Although significant improvement in overall survival has been achieved,death in adult populations under the age of 55 appears to have increased in the past decades.Although new classes of therapeutic strategies such as immunotherapy have emerged,their application is very limited in CRC so far.Microtubule(MT)inhibitors such as taxanes,are not generally successful in CRC.There may be some way to make MT inhibitors work effectively in CRC.One potential advantage that we can take to treat CRC may be the combination of optical techniques coupled to an endoscope or other fiber optics-based devices.A combination of optical devices and photo-activatable drugs may allow us to locally target advanced CRC cells with highly potent MT-targeting drugs.In this Editorial review,we would like to discuss the potential of optogenetic approaches in CRC management.展开更多
Over the years immunotherapy has demonstrably improved the field of cancer treatment.However,achieving long-term survival for colorectal cancer(CRC)patients remains a significant unmet need.Combination immunotherapies...Over the years immunotherapy has demonstrably improved the field of cancer treatment.However,achieving long-term survival for colorectal cancer(CRC)patients remains a significant unmet need.Combination immunotherapies incor-porating targeted drugs like MEK or multi-kinase inhibitors have offered some palliative benefit.Nevertheless,substantial gaps remain in the current therapeutic armamentarium for CRC.In recent years,there has been a surge of interest in exploring novel treatment strategies,including the application of light-activated drugs in conjunction with optical devices.This approach holds promise for achie-ving localized and targeted delivery of cytotoxic agents,such as microtubule-targeting drugs,directly to cancerous cells within the colon.展开更多
In order to understand the microtubule change of monocotyls stem-tip during mitosis, the arrangement, transformation of microtubule array and its relation with chromosome movement during mitosis were studied with free...In order to understand the microtubule change of monocotyls stem-tip during mitosis, the arrangement, transformation of microtubule array and its relation with chromosome movement during mitosis were studied with freezing microtome, indirect immunofluoreseenee, DAPI staining and fluorescence microscopy. The results showed that nucleolus was intact when the cortical microtubules formed; cortical microtubules were changed into phramoplast microtubules bands at mitosis prophase. When phramoplast microtubules came into being, nuclear membrane was ruptured and chromosome was arranged at the position of cell plate ; subsequently, phramoplast microtubules were changed into phragmoplast microtubules, phramoplast microtubules were shortening and microtubules on the sides of cell plate were increasing gradually, during this course sister ehromatid was separated by microtubules at cell plate and tract to the two poles, forming phragmoplast microtubules. Then the nucleolus of two daughter cells formed and separated in the end with the increase of cells numbers. Therefore, cell division orientation could be judged from the arrangement of cell microtubules in different periods in order to understand its growth status.展开更多
Tau, a primary component of microtubule-associated protein, promotes microtubule assembly and/or disassembly and maintains the stability of the microtubule structure. Although the importance of tau in neurodegenerativ...Tau, a primary component of microtubule-associated protein, promotes microtubule assembly and/or disassembly and maintains the stability of the microtubule structure. Although the importance of tau in neurodegenerative diseases has been well demonstrated, wheth- er tau is involved in peripheral nerve regeneration remains unknown. In the current study, we obtained sciatic nerve tissue from adult rats 0, 1, 4, 7, and 14 days after sciatic nerve crush and examined tau mRNA and protein expression levels and the location of tau in the sciatic nerve following peripheral nerve injury. The results from our quantitative reverse transcription polymerase chain reaction analysis showed that compared with the uninjured control sciatic nerve, mRNA expression levels for both tau and tau tubulin kinase 1, a serine/ threonine kinase that regulates tau phosphorylation, were decreased following peripheral nerve injury. Our western blot assay results suggested that the protein expression levels of tau and phosphorylated tau initially decreased 1 day post nerve injury but then gradually increased. The results of our immunohistochemical labeling showed that the location of tau protein was not altered by nerve injury. Thus, these results showed that the expression of tau was changed following sciatic nerve crush, suggesting that tau may be involved in periph- eral nerve repair and regeneration.展开更多
AIM To investigate the interaction of Zot withmicrotubule.METHODS Zot affinity column was applied topurify Zot-binding protein（s）from crudeintestinal cell lysates.After incubation at roomtemperature,the column was w...AIM To investigate the interaction of Zot withmicrotubule.METHODS Zot affinity column was applied topurify Zot-binding protein（s）from crudeintestinal cell lysates.After incubation at roomtemperature,the column was washed and theproteins bound to the Zot affinity column wereeluted by step gradient with NaCl(0.3 mol·L<sup>-1</sup>-0.5mol·L<sup>-1</sup>).The fractions were subjected to6.0%-15.0%（w/v）gradient SDS-PAGE andthen transferred to PVDF membrane for N-terminal sequencing.Purified Zot and tauprotein were blotted by using anti-Zot or anti-tauantibodies.Finally,purified Zot was tested in anin vitro tubulin binding assay.RESULTS Fractions from Zot affinity columnyielded two protein bands with a Mr of 60 kU and45kU respectively.The N-terminal sequence ofthe 60 kU band resulted identical to β-tubulin.Zot also cross-reacts with anti-tau antibodies.Inthe in vitro tubulin binding assay,Zot co-precipitate with Mt,further suggesting that Zotpossesses tubulin-binding properties.CONCLUSION Taken together,these resultssuggest that Zot regulates the permeability ofintestinal tight junctions by binding tointracellular Mt,with the subsequent activationof the intracellular signaling leading to thepermeabilization of intercellular tight junctions.展开更多
Fidgetin,a microtubule-severing enzyme,regulates neurite outgrowth,axonal regeneration,and cell migration by trimming off the labile domain of microtubule polymers.Because maintenance of the microtubule labile domain ...Fidgetin,a microtubule-severing enzyme,regulates neurite outgrowth,axonal regeneration,and cell migration by trimming off the labile domain of microtubule polymers.Because maintenance of the microtubule labile domain is essential for axon initiation,elongation,and navigation,it is of interest to determine whether augmenting the microtubule labile domain via depletion of fidgetin serves as a therapeutic approach to promote axonal regrowth in spinal cord injury.In this study,we constructed rat models of spinal cord injury and sciatic nerve injury.Compared with spinal cord injury,we found that expression level of tyrosinated microtubules in the labile portion of microtubules continuously increased,whereas fidgetin decreased after peripheral nerve injury.Depletion of fidgetin enhanced axon regeneration after spinal cord injury,whereas expression level of end binding protein 3(EB3)markedly increased.Next,we performed RNA interference to knockdown EB3 or fidgetin.We found that deletion of EB3 did not change fidgetin expression.Conversely,deletion of fidgetin markedly increased expression of tyrosinated microtubules and EB3.Deletion of fidgetin increased the amount of EB3 at the end of neurites and thereby increased the level of tyrosinated microtubules.Finally,we deleted EB3 and overexpressed fidgetin.We found that fidgetin trimmed tyrosinated tubulins by interacting with EB3.When fidgetin was deleted,the labile portion of microtubules was elongated,and as a result the length of axons and number of axon branches were increased.These findings suggest that fidgetin can be used as a novel therapeutic target to promote axonal regeneration after spinal cord injury.Furthermore,they reveal an innovative mechanism by which fidgetin preferentially severs labile microtubules.展开更多
Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in...Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in the brains of 1-week and 6-week old fragile X mental retardation-1 （FmrI） knockout （KO） mice were investigated by immunohistochemistry, Western blot, and in situ hybridization, with the age-matched wild type mice （WT） as controls. Results The mean optical density （MOD） of MAP1B was significantly decreased in each brain region in KO6W compared with WT6W, whereas in KO1W, this decrease was only found in the hippocampus and cerebellum. MAP1B in 6-week mice was much less than that in 1-week mice of the same genotype. The results of Western blot and in situ hybridization showed that MAP1B protein and MAP1B mRNA were significantly decreased in the hippocampus of both KO1W and KO6W. Conclusion The decreased MAP1B protein and MAP1B mRNA in the Fmrl knockout mice indicate that FMRP may positively regulate the expression of MAP1B.展开更多
Histological changes that occur during microsporogenesis are documented in a gene-cytoplasmic male sterile rice ( Oryza saliva L.) line, Zhen Shan 97A, its maintainer line, Zhen Shan 97B, and the restorer line, Ce64 o...Histological changes that occur during microsporogenesis are documented in a gene-cytoplasmic male sterile rice ( Oryza saliva L.) line, Zhen Shan 97A, its maintainer line, Zhen Shan 97B, and the restorer line, Ce64 of a Mine hybrid rice production system. In the restorer line, Ce64, the developing microsporocytes have dense cytoplasm and a distinct set of circumferential microtubules around the nucleus. Successive cytokinesis results in the formation of tetrads. The microtubules within the cells of tetrads and microspores radiate from the surface of the nucleus towards the outer edge of the cytoplasm. Subsequent pollen development is normal. During the course of microspore formation tubulin speckles can be found in the cytoplasm. The general pattern of development and microtubule organization in the maintainer lined Zhen Shan 97B, is similar to Ce64, except that a few more tubulin speckles appear during microspore formation. In the case of the mate sterile line, Zhen Shan 97A, a number of abnormalities can be discerned during early microsporogenesis. These include vacuoles forming within the developing microsporocyte and faintly stained microtubules with no defined distribution pattern. Prominent tubulin speckles are common within the cytoplasm. For those microsporocytes that undergo meiosis, no defined organizational patterns of microtubules can be found within the tetrad. All microspores abort soon after. Abnormalities and defects in microtubule organization observed in Zhen Shan 97A showed that complex interactions between the cytoplasm and the nucleus began at very early stage of microsporocyte development.展开更多
Changes in the pattern of organization of microtubules in the meiotic stages of development of pollen (i.e. from pre-meiotic interphase to more or less metaphase I) of a normal (IR36) and a temperature/photoperiod sen...Changes in the pattern of organization of microtubules in the meiotic stages of development of pollen (i.e. from pre-meiotic interphase to more or less metaphase I) of a normal (IR36) and a temperature/photoperiod sensitive male sterile line (Peiai 64S) of rice were studied using immunofluorescence confocal microscopy. In IR36, from pre-meiotic interphase to metaphase I, the pattern of microtubule distribution in the meiocytes underwent a series of changes. Some new organizational patterns of microtubules (that have not been described before) were observed during microsporogenesis, including the existence of a broad band of perinuclear microtubules at the diakinesis stage of development. The pattern of microtubule distribution in the meiocytes of the male sterile line, Peiai 64S, was quite different front that seen in IR36. In Peiai 64S, the microtubules showed abnormal patterns of distribution from pre-meiotic interphase to metaphase I. For example the broad band of perinuclear microtubules seen at diakinesis in IR36 was much disorganized and loosened in Peiai 64S. The spindles formed were also very abnormal and different from the normal spindle. The appearance of abnormal microtubule distribution in the early stages of microsporogenesis may contribute to the malformation and ultimate abortion of pollen in Peiai 64S.展开更多
Microtubule arrays in prothalli large-vacuolated and meristematic dividing cells of the fern Dryopteris crassirhizoma Nakai were studied using Steedman's wax, indirect immunofluorescence labelling and confocal las...Microtubule arrays in prothalli large-vacuolated and meristematic dividing cells of the fern Dryopteris crassirhizoma Nakai were studied using Steedman's wax, indirect immunofluorescence labelling and confocal laser scanning microscopy. Results showed that the use of high paraformaldehyde concentration (8%) allowed good fixation of prothallus cells, which are characterized by numerous (meristematic cells) and big (large-vacuolated cells) vacuoles. Results also plead for the efficiency of Steedman's wax embedding method in: (1) avoiding excessive use of enzyme for digesting cell wall in the process of the microtubule cytoskeleton labelling, (2) minimizing the autofluorescence effect in cells through utilization of alcohol in sample dehydration, and (3) permitting a clear visualization of microtubule patterns during the cell mitosis. Steedman's wax, coupled with immunofluorescence labelling and confocal laser scanning microscopy techniques, allows a good investigation of cell division process in plants by using simple multicellular organisms such as fern prothalli.展开更多
Changes in the pattern of microtubule distribution and organization during megagametogenesis in the embryo sac of rice (Oryza sativa L. cv. IR36) were re-examined using a modified polyethylene glycol sectioning techni...Changes in the pattern of microtubule distribution and organization during megagametogenesis in the embryo sac of rice (Oryza sativa L. cv. IR36) were re-examined using a modified polyethylene glycol sectioning technique before immuno-fluorescence staining of microtubules. In the sectioned materials the pattern of distribution and structural organization of the microtubule cytoskeleton were quite well preserved. Fine details of the patterns of structural changes and re-organization of the microtubule cytoskeleton in the major stages of development during embryo sac megagametogenesis (viz. functional megaspore, uni-nucleate, 2-nucleate, 4-nucleate, 8-nucleate and mature stage) could be clearly observed and easily followed. Some new organizational patterns of microtubules associated with the probable movement and positioning of the polar nuclei were observed.展开更多
The reorientation of microtubules (MTS) in roots of Oryza sativa L. treated with 2,4_D was studied using confocal laser scanning microscopy. In the control (the roots were not treated with 2,4_D), different distributi...The reorientation of microtubules (MTS) in roots of Oryza sativa L. treated with 2,4_D was studied using confocal laser scanning microscopy. In the control (the roots were not treated with 2,4_D), different distribution patterns of MTS were observed in the different growth zones of root tips. MTS of the cortical cells were randomly aligned in the zone of cell division. They were transversely arranged in the cortical cells of the zone of cell elongation; and obliquely oriented in the root hair zone. After treatment with 2,4_D, MTS displayed distinct changes with reorientation in the cortex of the root tip coupling with the inhibition of root growth. MTS changed their orientation in the cortical cells of the zone of cell division from being randomly oriented to transversely oriented when incubated in 1 mg/L 2,4_D for 1 h. However, they were recovered and became randomly oriented when the roots were treated with 1 mg/L 2,4_D for 24 h. An array of MTS, which was different from that in the control, was observed in the cortical cells of the zone of cell elongation in the roots treated with 1 mg/L 2,4_D for 1 h. After treatment with 10 mg/L 2,4_D for 1 h, the MTS in the cortical cells of the zone of cell division became transversely reorientated, but the pattern of MTS distribution was different from that in the 1_hour treatment of 1 mg/L 2,4_D. MTS were hardly detected in the same type of cells when roots were submerged in 10 mg/L 2,4_D for 24 h. When roots were incubated in 10 mg/L 2,4_D for 1 h, the MTS of the cortical cells in the zone of cell elongation became randomly oriented, and much more randomly when the roots were treated for 24 h.展开更多
In most kinds of animal cells, the centrosome serves as the main microtubule organizing center （MTOC） that nucleates microtubule arrays throughout the cytoplasm to maintain cell structure, cell division and intracel...In most kinds of animal cells, the centrosome serves as the main microtubule organizing center （MTOC） that nucleates microtubule arrays throughout the cytoplasm to maintain cell structure, cell division and intracellular transport. Whereas in epithelial cells, non-centrosomal MTOCs are established in the apical domain for generating asymmetric microtubule fibers and cilia in epithelial cells for the organ morphogenesis during embryonic development. However, the mechanism by which MTOCs localize to the apical domain in epithelial cells remains largely unknown. Here, we show that Mid liplb has a close interaction with γ-tubulin protein, the central component of MTOC, and modulates lumen opening of the neural tube, gut, intestine, and kidney of zebrafish. Knockdown or dominant negative effect of Mid 1ip 1 b resulted in failure of lumen formation of the organs as aforementioned. Moreover, the non-centrosomal MTOCs were unable to orientate to the apical domain in Midliplb knockdown epithelial cells, and the centrosomal MTOCs were inaccurately placed in the apical domain, resulting in defective formation of asymmetric microtubules and misplacement of cilia in the apical domain. These data uncover a molecule that controls the proper localization of MTOCs in the apical domain in epithelial cells for organ morphogenesis during embryonic development.展开更多
Microtubule self-organization under mechanical and chemical regulations plays a central role in cytokinesis and cel-lular transportations.In plant-cells,the patterns or phases of cortical microtubules organizations ar...Microtubule self-organization under mechanical and chemical regulations plays a central role in cytokinesis and cel-lular transportations.In plant-cells,the patterns or phases of cortical microtubules organizations are the direct indicators of cell-phases.The dense nematic pattern of cortical microtubule array relies on the regulation of single microtubule dynamics with mechanical coupling to steric interaction among the self-organized microtubule crowds.Building upon previous mini-mal models,we investigate the effective microtubule width,microtubule catastrophe rate,and zippering angle as factors that regulate the self-organization of the dense nematic phase.We find that by incorporating the effective microtubule width,the transition from isotropic to the highly ordered nematic phase(NI phase)with extremely long microtubules will be gapped by another nematic phase which consists of relative short microtubules(NII phase).The NII phase in the gap grows wider with the increase of the microtubule width.We further illustrate that in the dense phase,the collision-induced catastrophe rate and an optimal zippering angle play an important role in controlling the order-disorder transition,as a result of the coupling between the collision events and ordering.Our study shows that the transition to dense microtubule array requires the cross-talk between single microtubule growth and mechanical interactions among microtubules in the active crowds.展开更多
Objective: To investigate the different inhibition effects of different sequential usages of microtubule depolymerization drug and polymerization drug on tumor cells. Methods: Three tumor cell lines including MCF-7, S...Objective: To investigate the different inhibition effects of different sequential usages of microtubule depolymerization drug and polymerization drug on tumor cells. Methods: Three tumor cell lines including MCF-7, SK-OV3, A549 were incubated with paclitaxel (PTX) and/or vinorelbine (NVB) of different concentrations. The cyto-toxicity was exam- ined by MTT test after incubating 72 h. According to different drugs and different sequences added to 96-well tissue culture plates, 5 groups were divided: PTX group (Group 1), NVB group (Group 2), PTX plus NVB group (Group 3), PTX first and NVB 4-h-later group (Group 4), and NVB first and PTX 4-h-later group (Group 5). Drug concentrations were 100% peak plasma concentration (PPC), 50% PPC, 25% PPC, 12.5% PPC, and 6. 25% PPC. Results: The inhibition effects on the three tumor cell lines in Group 5 were stronger than those in the other four groups (P < 0.01). And the inhibition effects in Group 4 were not stronger than those in Groups 1, 2 or 3 (P > 0.1). Conclusion: Using microtubule depolymerization drug first and then using microtubule polymerization drug has synergic inhibition effect on tumor cells.展开更多
Allicin, a major biologically active component of garlic, is produced from its inactive precursor aUiin by the enzyme alliinase. In this study, we investigated its effects on human nasopharyngeal carcinoma KB cells. A...Allicin, a major biologically active component of garlic, is produced from its inactive precursor aUiin by the enzyme alliinase. In this study, we investigated its effects on human nasopharyngeal carcinoma KB cells. After incubation for 48 h, allicin inhibited the growth of KB cells in a concentration-dependent manner with an IC50 value of （2.2±0.2） μg/mL. Incubation with allicin for 48 h caused a concentration-dependent induction of apoptosis in the concentration range of （16-48） μg/mL, and the induction of apoptosis was confirmed by the changes of mitochondrial membrane potential, F-actin contents and nuclear condensation in KB cells. Moreover, allicin concentration-dependently arrested KB cells at the S-phase of the cell cycle in the range of （16-48） μg/mL. In addition, treatment with the compound caused concentration-dependent disassembly of microtubule cytoskeleton in KB cells, which is similar to the effect of colchicine, a well-known microtubule destabilizing agent. We concluded that the abilities of allicin to inhibit the proliferation of KB cells probably relate to its apoptosis induction, cell cycle arrest and microtubule destabilizing properties.展开更多
基金supported by the National Nature Science Foundation of China,No.32101020(to JL)the Natural Science Foundation of Shandong Province,Nos.ZR2020MC071(to JL),ZR2023MH327(to HZ)+1 种基金the Integrated Project of Major Research Plan of National Natural Science Foundation of China,No.92249303(to PL)the Natural Science Foundation of Qingdao,No.23-2-1-193-zyyd-jch(to HZ)。
文摘The protein connector enhancer of kinase suppressor of Ras 2(CNKSR2),present in both the postsynaptic density and cytoplasm of neurons,is a scaffolding protein with several protein-binding domains.Variants of the CNKSR2 gene have been implicated in neurodevelopmental disorders,particularly intellectual disability,although the precise mechanism involved has not yet been fully understood.Research has demonstrated that CNKSR2 plays a role in facilitating the localization of postsynaptic density protein complexes to the membrane,thereby influencing synaptic signaling and the morphogenesis of dendritic spines.However,the function of CNKSR2 in the cytoplasm remains to be elucidated.In this study,we used immunoprecipitation and high-resolution liquid chromatography-mass spectrometry to identify the interactors of CNKSR2.Through a combination of bioinformatic analysis and cytological experiments,we found that the CNKSR2 interactors were significantly enriched in the proteome of the centrosome.We also showed that CNKSR2 interacted with the microtubule protein DYNC1H1 and with the centrosome marker CEP290.Subsequent colocalization analysis confirmed the centrosomal localization of CNKSR2.When we downregulated CNKSR2 expression in mouse neuroblastoma cells(Neuro 2A),we observed significant changes in the expression of numerous centrosomal genes.This manipulation also affected centrosome-related functions,including cell size and shape,cell proliferation,and motility.Furthermore,we found that CNKSR2 interactors were highly enriched in de novo variants associated with intellectual disability and autism spectrum disorder.Our findings establish a connection between CNKSR2 and the centrosome,and offer new insights into the underlying mechanisms of neurodevelopmental disorders.
基金supported by the Koeln Fortune Program/Faculty of Medicine,University of Cologne,the Alzheimer Forschung Initiative e.V.(grant#22039,to HZ)open-access funding from the DFG/GRC issued to the University of CologneAlzheimer Forschung Initiative e.V.for Open Access Publishing(a publication grant#P2401,to MAAK)。
文摘TAU is a microtubule-associated protein that promotes microtubule assembly and stability in the axon.TAU is missorted and aggregated in an array of diseases known as tauopathies.Microtubules are essential for neuronal function and regulated via a complex set of post-translational modifications,changes of which affect microtubule stability and dynamics,microtubule interaction with other proteins and cellular structures,and mediate recruitment of microtubule-severing enzymes.As impairment of microtubule dynamics causes neuronal dysfunction,we hypothesize cognitive impairment in human disease to be impacted by impairment of microtubule dynamics.We therefore aimed to study the effects of a disease-causing mutation of TAU(P301L)on the levels and localization of microtubule post-translational modifications indicative of microtubule stability and dynamics,to assess whether P301L-TAU causes stability-changing modifications to microtubules.To investigate TAU localization,phosphorylation,and effects on tubulin post-translational modifications,we expressed wild-type or P301L-TAU in human MAPT-KO induced pluripotent stem cell-derived neurons(i Neurons)and studied TAU in neurons in the hippocampus of mice transgenic for human P301L-TAU(p R5 mice).Human neurons expressing the longest TAU isoform(2N4R)with the P301L mutation showed increased TAU phosphorylation at the AT8,but not the p-Ser-262 epitope,and increased polyglutamylation and acetylation of microtubules compared with endogenous TAU-expressing neurons.P301L-TAU showed pronounced somatodendritic presence,but also successful axonal enrichment and a similar axodendritic distribution comparable to exogenously expressed 2N4R-wildtype-TAU.P301L-TAU-expressing hippocampal neurons in transgenic mice showed prominent missorting and tauopathy-typical AT8-phosphorylation of TAU and increased polyglutamylation,but reduced acetylation,of microtubules compared with non-transgenic littermates.In sum,P301L-TAU results in changes in microtubule PTMs,suggestive of impairment of microtubule stability.This is accompanied by missorting and aggregation of TAU in mice but not in i Neurons.Microtubule PTMs/impairment may be of key importance in tauopathies.
基金supported by the National Natural Science Foundation of China,No.32071033(to MT)Basic and Applied Basic Research Foundation of Guangdong Province,Nos.2023A1515010140(to MT),2022A1515140169(to MT),2022A1515111096(to ZC)+3 种基金Science and Technology Project of Guangzhou,Nos.202201010015(to YL),2023A03J0790(to TJ)Basic and Applied Basic Research Foundation of Guangzhou,No.2023A04J1285(to ZC)Medical Research Foundation of Guangdong Province,No.A2023147(to ZC)Health Science and Technology Project of Guangzhou,No.20221A011039(to TJ)。
文摘Post-translational modification of spastin enables precise spatiotemporal control of its microtubule severing activity.However,the detailed mechanism by which spastin turnover is regulated in the context of neurite outgrowth remains unknown.Here,we found that spastin interacted with ubiquitin and was significantly degraded by K48-mediated poly-ubiquitination.Cullin3 facilitated spastin degradation and ubiquitination.RING-box protein 1,but not RING-box protein 2,acted synergistically with Cullin3 protein to regulate spastin degradation.Overexpression of Culin3 or BRX1 markedly suppressed spastin expression,and inhibited spastin-mediated microtubule severing and promotion of neurite outgrowth.Moreover,USP14 interacted directly with spastin to mediate its deubiquitination.USP14 overexpression significantly increased spastin expression and suppressed its ubiquitination and degradation.Although co-expression of spastin and USP14 did not enhance microtubule severing,it did increase neurite length in hippocampal neurons.Taken together,these findings elucidate the intricate regulatory mechanisms of spastin turnover,highlighting the roles of the Cullin-3–Ring E3 ubiquitin ligase complex and USP14 in orchestrating its ubiquitination and degradation.The dynamic interplay between these factors governs spastin stability and function,ultimately influencing microtubule dynamics and neuronal morphology.These insights shed light on potential therapeutic targets for neurodegenerative disorders associated with spastin defects.
基金Project supported by the Natural Science Foundation of Hebei Province of China (Grant No. A2023202010)。
文摘Microtubules are one kind of cytoskeleton that is ubiquitous in eukaryotic cells and is essential for various biological processes, such as intracellular transport, maintenance of cell morphology and cell division. Microtubules are dynamic structures and the basic unit of microtubules is the heterodimer composed of α-tubulin and β-tubulin. The biological function of microtubules is based on their dynamic polymerization and depolymerization. However, the nucleotide-dependent depolymerization mechanism of microtubules is still unclear. The dynamic instability of microtubules is determined by the interactions between tubulins. In this work, the interactions between tubulins in the microtubule lattice(GDP at the interdimer interface) and in the special GTP-cap structure(GTP at the interdimer interface) are systematically investigated using all-atom molecular dynamics simulation method. The analysis of the tubulin–tubulin and nucleotide–tubulin interactions and binding free energy at different interfaces of microtubule shows that the hydrolysis of GTP can affect the longitudinal interaction between α-tubulin and β-tubulin at the interdimer interface and the lateral interaction between β-tubulins. In particular, the displacement of M loop of β-tubulin induced by GTP hydrolysis weakens the lateral interaction betweenβ-tubulins. Based on these results, a nucleotide-dependent depolymerization mechanism of microtubule induced by GTP hydrolysis is proposed.
文摘Colorectal cancer(CRC)has remained the second and the third leading cause of cancer-related death worldwide and in the United States,respectively.Although significant improvement in overall survival has been achieved,death in adult populations under the age of 55 appears to have increased in the past decades.Although new classes of therapeutic strategies such as immunotherapy have emerged,their application is very limited in CRC so far.Microtubule(MT)inhibitors such as taxanes,are not generally successful in CRC.There may be some way to make MT inhibitors work effectively in CRC.One potential advantage that we can take to treat CRC may be the combination of optical techniques coupled to an endoscope or other fiber optics-based devices.A combination of optical devices and photo-activatable drugs may allow us to locally target advanced CRC cells with highly potent MT-targeting drugs.In this Editorial review,we would like to discuss the potential of optogenetic approaches in CRC management.
文摘Over the years immunotherapy has demonstrably improved the field of cancer treatment.However,achieving long-term survival for colorectal cancer(CRC)patients remains a significant unmet need.Combination immunotherapies incor-porating targeted drugs like MEK or multi-kinase inhibitors have offered some palliative benefit.Nevertheless,substantial gaps remain in the current therapeutic armamentarium for CRC.In recent years,there has been a surge of interest in exploring novel treatment strategies,including the application of light-activated drugs in conjunction with optical devices.This approach holds promise for achie-ving localized and targeted delivery of cytotoxic agents,such as microtubule-targeting drugs,directly to cancerous cells within the colon.
基金Supported by the National Natural Science Foundation of China(30060038)~~
文摘In order to understand the microtubule change of monocotyls stem-tip during mitosis, the arrangement, transformation of microtubule array and its relation with chromosome movement during mitosis were studied with freezing microtome, indirect immunofluoreseenee, DAPI staining and fluorescence microscopy. The results showed that nucleolus was intact when the cortical microtubules formed; cortical microtubules were changed into phramoplast microtubules bands at mitosis prophase. When phramoplast microtubules came into being, nuclear membrane was ruptured and chromosome was arranged at the position of cell plate ; subsequently, phramoplast microtubules were changed into phragmoplast microtubules, phramoplast microtubules were shortening and microtubules on the sides of cell plate were increasing gradually, during this course sister ehromatid was separated by microtubules at cell plate and tract to the two poles, forming phragmoplast microtubules. Then the nucleolus of two daughter cells formed and separated in the end with the increase of cells numbers. Therefore, cell division orientation could be judged from the arrangement of cell microtubules in different periods in order to understand its growth status.
基金supported by the National Natural Science Foundation of China,No.81130080,31300942the National Key Basic Research Program of China(973 Program)+5 种基金No.2014CB542202the Natural Science Foundation of Jiangsu Province,China,No.BK20150409the Natural Science Foundation of Jiangsu Higher Education Institutions of China,No.15KJB180013the Scientific Research Foundation of Nantong University of China,No.14R29the Natural Science Foundation of Nantong City in China,No.MS12015043the Priority Academic Program Development of Jiangsu Higher Education Institutions of China
文摘Tau, a primary component of microtubule-associated protein, promotes microtubule assembly and/or disassembly and maintains the stability of the microtubule structure. Although the importance of tau in neurodegenerative diseases has been well demonstrated, wheth- er tau is involved in peripheral nerve regeneration remains unknown. In the current study, we obtained sciatic nerve tissue from adult rats 0, 1, 4, 7, and 14 days after sciatic nerve crush and examined tau mRNA and protein expression levels and the location of tau in the sciatic nerve following peripheral nerve injury. The results from our quantitative reverse transcription polymerase chain reaction analysis showed that compared with the uninjured control sciatic nerve, mRNA expression levels for both tau and tau tubulin kinase 1, a serine/ threonine kinase that regulates tau phosphorylation, were decreased following peripheral nerve injury. Our western blot assay results suggested that the protein expression levels of tau and phosphorylated tau initially decreased 1 day post nerve injury but then gradually increased. The results of our immunohistochemical labeling showed that the location of tau protein was not altered by nerve injury. Thus, these results showed that the expression of tau was changed following sciatic nerve crush, suggesting that tau may be involved in periph- eral nerve repair and regeneration.
文摘AIM To investigate the interaction of Zot withmicrotubule.METHODS Zot affinity column was applied topurify Zot-binding protein（s）from crudeintestinal cell lysates.After incubation at roomtemperature,the column was washed and theproteins bound to the Zot affinity column wereeluted by step gradient with NaCl(0.3 mol·L<sup>-1</sup>-0.5mol·L<sup>-1</sup>).The fractions were subjected to6.0%-15.0%（w/v）gradient SDS-PAGE andthen transferred to PVDF membrane for N-terminal sequencing.Purified Zot and tauprotein were blotted by using anti-Zot or anti-tauantibodies.Finally,purified Zot was tested in anin vitro tubulin binding assay.RESULTS Fractions from Zot affinity columnyielded two protein bands with a Mr of 60 kU and45kU respectively.The N-terminal sequence ofthe 60 kU band resulted identical to β-tubulin.Zot also cross-reacts with anti-tau antibodies.Inthe in vitro tubulin binding assay,Zot co-precipitate with Mt,further suggesting that Zotpossesses tubulin-binding properties.CONCLUSION Taken together,these resultssuggest that Zot regulates the permeability ofintestinal tight junctions by binding tointracellular Mt,with the subsequent activationof the intracellular signaling leading to thepermeabilization of intercellular tight junctions.
基金the National Natural Science Foundation of China,Nos.32070725(to ML),82001295(to RHW),31970412(to YL)the Priority Academic Program Development(PAPD)of Jiangsu Higher Education Institutions。
文摘Fidgetin,a microtubule-severing enzyme,regulates neurite outgrowth,axonal regeneration,and cell migration by trimming off the labile domain of microtubule polymers.Because maintenance of the microtubule labile domain is essential for axon initiation,elongation,and navigation,it is of interest to determine whether augmenting the microtubule labile domain via depletion of fidgetin serves as a therapeutic approach to promote axonal regrowth in spinal cord injury.In this study,we constructed rat models of spinal cord injury and sciatic nerve injury.Compared with spinal cord injury,we found that expression level of tyrosinated microtubules in the labile portion of microtubules continuously increased,whereas fidgetin decreased after peripheral nerve injury.Depletion of fidgetin enhanced axon regeneration after spinal cord injury,whereas expression level of end binding protein 3(EB3)markedly increased.Next,we performed RNA interference to knockdown EB3 or fidgetin.We found that deletion of EB3 did not change fidgetin expression.Conversely,deletion of fidgetin markedly increased expression of tyrosinated microtubules and EB3.Deletion of fidgetin increased the amount of EB3 at the end of neurites and thereby increased the level of tyrosinated microtubules.Finally,we deleted EB3 and overexpressed fidgetin.We found that fidgetin trimmed tyrosinated tubulins by interacting with EB3.When fidgetin was deleted,the labile portion of microtubules was elongated,and as a result the length of axons and number of axon branches were increased.These findings suggest that fidgetin can be used as a novel therapeutic target to promote axonal regeneration after spinal cord injury.Furthermore,they reveal an innovative mechanism by which fidgetin preferentially severs labile microtubules.
文摘Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in the brains of 1-week and 6-week old fragile X mental retardation-1 （FmrI） knockout （KO） mice were investigated by immunohistochemistry, Western blot, and in situ hybridization, with the age-matched wild type mice （WT） as controls. Results The mean optical density （MOD） of MAP1B was significantly decreased in each brain region in KO6W compared with WT6W, whereas in KO1W, this decrease was only found in the hippocampus and cerebellum. MAP1B in 6-week mice was much less than that in 1-week mice of the same genotype. The results of Western blot and in situ hybridization showed that MAP1B protein and MAP1B mRNA were significantly decreased in the hippocampus of both KO1W and KO6W. Conclusion The decreased MAP1B protein and MAP1B mRNA in the Fmrl knockout mice indicate that FMRP may positively regulate the expression of MAP1B.
文摘Histological changes that occur during microsporogenesis are documented in a gene-cytoplasmic male sterile rice ( Oryza saliva L.) line, Zhen Shan 97A, its maintainer line, Zhen Shan 97B, and the restorer line, Ce64 of a Mine hybrid rice production system. In the restorer line, Ce64, the developing microsporocytes have dense cytoplasm and a distinct set of circumferential microtubules around the nucleus. Successive cytokinesis results in the formation of tetrads. The microtubules within the cells of tetrads and microspores radiate from the surface of the nucleus towards the outer edge of the cytoplasm. Subsequent pollen development is normal. During the course of microspore formation tubulin speckles can be found in the cytoplasm. The general pattern of development and microtubule organization in the maintainer lined Zhen Shan 97B, is similar to Ce64, except that a few more tubulin speckles appear during microspore formation. In the case of the mate sterile line, Zhen Shan 97A, a number of abnormalities can be discerned during early microsporogenesis. These include vacuoles forming within the developing microsporocyte and faintly stained microtubules with no defined distribution pattern. Prominent tubulin speckles are common within the cytoplasm. For those microsporocytes that undergo meiosis, no defined organizational patterns of microtubules can be found within the tetrad. All microspores abort soon after. Abnormalities and defects in microtubule organization observed in Zhen Shan 97A showed that complex interactions between the cytoplasm and the nucleus began at very early stage of microsporocyte development.
文摘Changes in the pattern of organization of microtubules in the meiotic stages of development of pollen (i.e. from pre-meiotic interphase to more or less metaphase I) of a normal (IR36) and a temperature/photoperiod sensitive male sterile line (Peiai 64S) of rice were studied using immunofluorescence confocal microscopy. In IR36, from pre-meiotic interphase to metaphase I, the pattern of microtubule distribution in the meiocytes underwent a series of changes. Some new organizational patterns of microtubules (that have not been described before) were observed during microsporogenesis, including the existence of a broad band of perinuclear microtubules at the diakinesis stage of development. The pattern of microtubule distribution in the meiocytes of the male sterile line, Peiai 64S, was quite different front that seen in IR36. In Peiai 64S, the microtubules showed abnormal patterns of distribution from pre-meiotic interphase to metaphase I. For example the broad band of perinuclear microtubules seen at diakinesis in IR36 was much disorganized and loosened in Peiai 64S. The spindles formed were also very abnormal and different from the normal spindle. The appearance of abnormal microtubule distribution in the early stages of microsporogenesis may contribute to the malformation and ultimate abortion of pollen in Peiai 64S.
文摘Microtubule arrays in prothalli large-vacuolated and meristematic dividing cells of the fern Dryopteris crassirhizoma Nakai were studied using Steedman's wax, indirect immunofluorescence labelling and confocal laser scanning microscopy. Results showed that the use of high paraformaldehyde concentration (8%) allowed good fixation of prothallus cells, which are characterized by numerous (meristematic cells) and big (large-vacuolated cells) vacuoles. Results also plead for the efficiency of Steedman's wax embedding method in: (1) avoiding excessive use of enzyme for digesting cell wall in the process of the microtubule cytoskeleton labelling, (2) minimizing the autofluorescence effect in cells through utilization of alcohol in sample dehydration, and (3) permitting a clear visualization of microtubule patterns during the cell mitosis. Steedman's wax, coupled with immunofluorescence labelling and confocal laser scanning microscopy techniques, allows a good investigation of cell division process in plants by using simple multicellular organisms such as fern prothalli.
文摘Changes in the pattern of microtubule distribution and organization during megagametogenesis in the embryo sac of rice (Oryza sativa L. cv. IR36) were re-examined using a modified polyethylene glycol sectioning technique before immuno-fluorescence staining of microtubules. In the sectioned materials the pattern of distribution and structural organization of the microtubule cytoskeleton were quite well preserved. Fine details of the patterns of structural changes and re-organization of the microtubule cytoskeleton in the major stages of development during embryo sac megagametogenesis (viz. functional megaspore, uni-nucleate, 2-nucleate, 4-nucleate, 8-nucleate and mature stage) could be clearly observed and easily followed. Some new organizational patterns of microtubules associated with the probable movement and positioning of the polar nuclei were observed.
文摘The reorientation of microtubules (MTS) in roots of Oryza sativa L. treated with 2,4_D was studied using confocal laser scanning microscopy. In the control (the roots were not treated with 2,4_D), different distribution patterns of MTS were observed in the different growth zones of root tips. MTS of the cortical cells were randomly aligned in the zone of cell division. They were transversely arranged in the cortical cells of the zone of cell elongation; and obliquely oriented in the root hair zone. After treatment with 2,4_D, MTS displayed distinct changes with reorientation in the cortex of the root tip coupling with the inhibition of root growth. MTS changed their orientation in the cortical cells of the zone of cell division from being randomly oriented to transversely oriented when incubated in 1 mg/L 2,4_D for 1 h. However, they were recovered and became randomly oriented when the roots were treated with 1 mg/L 2,4_D for 24 h. An array of MTS, which was different from that in the control, was observed in the cortical cells of the zone of cell elongation in the roots treated with 1 mg/L 2,4_D for 1 h. After treatment with 10 mg/L 2,4_D for 1 h, the MTS in the cortical cells of the zone of cell division became transversely reorientated, but the pattern of MTS distribution was different from that in the 1_hour treatment of 1 mg/L 2,4_D. MTS were hardly detected in the same type of cells when roots were submerged in 10 mg/L 2,4_D for 24 h. When roots were incubated in 10 mg/L 2,4_D for 1 h, the MTS of the cortical cells in the zone of cell elongation became randomly oriented, and much more randomly when the roots were treated for 24 h.
基金supported by the National Basic Research Program of China (to X.M.,2015CB942800)the Nature Science Foundation of China (to H. X. 31671502)
文摘In most kinds of animal cells, the centrosome serves as the main microtubule organizing center （MTOC） that nucleates microtubule arrays throughout the cytoplasm to maintain cell structure, cell division and intracellular transport. Whereas in epithelial cells, non-centrosomal MTOCs are established in the apical domain for generating asymmetric microtubule fibers and cilia in epithelial cells for the organ morphogenesis during embryonic development. However, the mechanism by which MTOCs localize to the apical domain in epithelial cells remains largely unknown. Here, we show that Mid liplb has a close interaction with γ-tubulin protein, the central component of MTOC, and modulates lumen opening of the neural tube, gut, intestine, and kidney of zebrafish. Knockdown or dominant negative effect of Mid 1ip 1 b resulted in failure of lumen formation of the organs as aforementioned. Moreover, the non-centrosomal MTOCs were unable to orientate to the apical domain in Midliplb knockdown epithelial cells, and the centrosomal MTOCs were inaccurately placed in the apical domain, resulting in defective formation of asymmetric microtubules and misplacement of cilia in the apical domain. These data uncover a molecule that controls the proper localization of MTOCs in the apical domain in epithelial cells for organ morphogenesis during embryonic development.
基金Project supported by the National Natural Science Foundation of China(Grant Nos.11474155,11774147,11674236,and 11922506)。
文摘Microtubule self-organization under mechanical and chemical regulations plays a central role in cytokinesis and cel-lular transportations.In plant-cells,the patterns or phases of cortical microtubules organizations are the direct indicators of cell-phases.The dense nematic pattern of cortical microtubule array relies on the regulation of single microtubule dynamics with mechanical coupling to steric interaction among the self-organized microtubule crowds.Building upon previous mini-mal models,we investigate the effective microtubule width,microtubule catastrophe rate,and zippering angle as factors that regulate the self-organization of the dense nematic phase.We find that by incorporating the effective microtubule width,the transition from isotropic to the highly ordered nematic phase(NI phase)with extremely long microtubules will be gapped by another nematic phase which consists of relative short microtubules(NII phase).The NII phase in the gap grows wider with the increase of the microtubule width.We further illustrate that in the dense phase,the collision-induced catastrophe rate and an optimal zippering angle play an important role in controlling the order-disorder transition,as a result of the coupling between the collision events and ordering.Our study shows that the transition to dense microtubule array requires the cross-talk between single microtubule growth and mechanical interactions among microtubules in the active crowds.
文摘Objective: To investigate the different inhibition effects of different sequential usages of microtubule depolymerization drug and polymerization drug on tumor cells. Methods: Three tumor cell lines including MCF-7, SK-OV3, A549 were incubated with paclitaxel (PTX) and/or vinorelbine (NVB) of different concentrations. The cyto-toxicity was exam- ined by MTT test after incubating 72 h. According to different drugs and different sequences added to 96-well tissue culture plates, 5 groups were divided: PTX group (Group 1), NVB group (Group 2), PTX plus NVB group (Group 3), PTX first and NVB 4-h-later group (Group 4), and NVB first and PTX 4-h-later group (Group 5). Drug concentrations were 100% peak plasma concentration (PPC), 50% PPC, 25% PPC, 12.5% PPC, and 6. 25% PPC. Results: The inhibition effects on the three tumor cell lines in Group 5 were stronger than those in the other four groups (P < 0.01). And the inhibition effects in Group 4 were not stronger than those in Groups 1, 2 or 3 (P > 0.1). Conclusion: Using microtubule depolymerization drug first and then using microtubule polymerization drug has synergic inhibition effect on tumor cells.
基金Grant of Modernization and Industrization of Weiur Medicine (Grant No. 2007BAI30B02-4)the grant from State Key Laboratory of Natural and Biomimetic Drugs, Peking University
文摘Allicin, a major biologically active component of garlic, is produced from its inactive precursor aUiin by the enzyme alliinase. In this study, we investigated its effects on human nasopharyngeal carcinoma KB cells. After incubation for 48 h, allicin inhibited the growth of KB cells in a concentration-dependent manner with an IC50 value of （2.2±0.2） μg/mL. Incubation with allicin for 48 h caused a concentration-dependent induction of apoptosis in the concentration range of （16-48） μg/mL, and the induction of apoptosis was confirmed by the changes of mitochondrial membrane potential, F-actin contents and nuclear condensation in KB cells. Moreover, allicin concentration-dependently arrested KB cells at the S-phase of the cell cycle in the range of （16-48） μg/mL. In addition, treatment with the compound caused concentration-dependent disassembly of microtubule cytoskeleton in KB cells, which is similar to the effect of colchicine, a well-known microtubule destabilizing agent. We concluded that the abilities of allicin to inhibit the proliferation of KB cells probably relate to its apoptosis induction, cell cycle arrest and microtubule destabilizing properties.
