TY - JOUR. T1 - Dose-dependent effect of nocodazole on endothelial cell cytoskeleton. AU - Smurova, K. M.. AU - Birukova, A. A.. AU - Verin, A. D.. AU - Alieva, Irina B.. PY - 2008/6/1. Y1 - 2008/6/1. N2 - The endothelium lining the inner surface of blood vessels fulfils an important barrier function and specifically, it controls vascular membrane permeability as well as nutrient and metabolite exchange in circulating blood and tissue fluids. Disturbances in vascular endothelium barrier function (vascular endothelium dysfunction) are coupled to cytoskeleton rearrangements, actomyosin contractility, and as a consequence, formation of paracellular gaps between endothelial cells. Microtubules constitute the first effector link in the reaction cascade resulting in vascular endothelium dysfunction. Increased vascular permeability associated with many human diseases is also manifested as a side effect in anticancer mitosis-blocking therapy. The aim of this study was to examine the possibility of ...

Here we investigated microtubule dynamics and function during neutrophil migration in vivo. In accordance with reported in vitro findings, we found that microtubule disruption impairs neutrophil directed migration to wounds but induces cell motility and polarity in a PI(3)K-independent manner. We also found that Rho mediates nocodazole-induced neutrophil motility. These findings are consistent with previous findings in vitro, but further provide a physiological context for neutrophil migration. In addition, we elucidated two previously unidentified roles of microtubules during neutrophil motility in vivo. First, microtubule arrays nucleate in front of the nucleus and mainly radiate towards the uropod. Second, Rac is activated by microtubule depolymerization in primary human neutrophils and is necessary for nocodazole-induced neutrophil motility in vivo.. Leukocytes are often described as having the microtubule cytoskeleton and MTOC behind the nucleus at the uropod (Friedl and Weigelin, 2008; ...

CLIP-170 staining after nocodazole treatment. CLIP-170 appears in green and chromosomes in red. 10-15 optical sections were merged for the final images. Oocyt

A previous report has shown that microtubule depolymerization by nocodazole treatment arrests cell cycle extracts in M phase if containing very high densities of sperm nuclei. The use of MKP-1 further suggested that MAP kinase, the most likely target of MKP-1, may have a role in the spindle assembly checkpoint (20). However, whether classical MAP kinase is involved in the mitotic arrest remains to be determined. To address this question, we used MAP kinase-depleted extracts. We monitored the level of MPF as histone H1 kinase activity in the extracts supplemented with 9,000 sperm nuclei/μl. In the absence of nocodazole, the extracts returned to interphase after the first M phase, irrespective of whether MAP kinase was depleted or not (Fig. 3,A). In the presence of nocodazole, mock-treated extracts were arrested at the first M phase and the high MPF activity was sustained (Fig. 3,B, mock), whereas MAP kinase-depleted extracts returned to interphase after the first M phase (Fig. 3,B, αMAPK). In ...

Lasers have made the 21st century even more technologically advanced with the are not role models, implementation at the cellular level. How To. Confocal microscopy alternatively to ultraviolent mercury lamps, are higher in wavelength and are more detailed in illumination in regards to shapes of sport stars role anymore, organelles (Katula). In this experiment, using confocal microscopy, two organelles were visualized to history understand the drug effect of nocodazole on NIH3T3 cells. Using two fluorescent stains, Mitotracker Red (accumulates in mitochondria) and role Hoechst 33342 (accumulates in nuclei) were used to test what microtubules would look like resulting from the drug. Nocodazole is a drug that penetrates the microtubules to where it can be synchronized. Ap World Essay 2007. (Karbowski). Microtubules are made up of polarized tubulin that are important for cytoskeletal rigidity, mitotic division, and act as a branch in sport role models anymore transporting molecules to other parts ...

Figure 1. High‐throughput siRNA screen identifies kinases that regulate the ionizing‐radiation‐induced G2 checkpoint. (A) Schematic overview of the robot‐automated high‐throughput siRNA screen. p53 dominant‐negative U2OS (U2OS p53dneg) cells were reverse transfected and, after incubation for 2 days, treated with 6 Gy of IR and, 2 h later, with nocodazole for 8 h and subsequently stained with H3Ser10p antibody to stain mitotic cells, and Hoechst to stain the nuclei. (B) Flow cytometric analysis of U2OS p53dneg cells. Where indicated, cells were treated with IR (6 Gy) and 2 h later with nocodazole for 8 h. Cells were collected and stained with phospho H3ser10 (H3Ser10p) antibody and propidium iodide (PI). (C) CHK1 depletion abrogates the G2 checkpoint. U2OS p53dneg cells were transfected with CHK1 siRNA and treated with IR (6 Gy) and 2 h later with nocodazole for 8 h. Top panel: immunofluorescence microscopy images obtained from the robot‐automated screen. Bottom panel: cells were ...

Author(s): Breitfeld, PP; McKinnon, WC; Mostov, KE | Abstract: A polarized cell, to maintain distinct basolateral and apical membrane domains, must tightly regulate vesicular traffic terminating at either membrane domain. In this study we have examined the extent to which microtubules regulate such traffic in polarized cells. Using the polymeric immunoglobulin receptor expressed in polarized MDCK cells, we have examined the effects of nocodazole, a microtubule-disrupting agent, on three pathways that deliver proteins to the apical surface and two pathways that deliver proteins to the basolateral surface. The biosynthetic and transcytotic pathways to the apical surface are dramatically altered by nocodazole in that a portion of the protein traffic on each of these two pathways is misdirected to the basolateral surface. The apical recycling pathway is slowed in the presence of nocodazole but targeting is not disrupted. In contrast, the biosynthetic and recycling pathways to the basolateral surface are

Hypoparathyroidism, mental retardation and facial dysmorphism (HRD) is a fatal developmental disease caused by mutations in tubulin-specific chaperone E (TBCE). A mouse Tbce mutation causes progressive motor neuronopathy. To dissect the functions of TBCE and the pathogenesis of HRD, mutations were generated in Drosophila tbce, and its expression was manipulated in a tissue-specific manner. Drosophila tbce nulls are embryonic lethal. Tissue-specific knockdown and overexpression of tbce in the neuromusculature resulted in disrupted and increased microtubules, respectively. Alterations in TBCE expression also affected neuromuscular synapses. Genetic analyses revealed an antagonistic interaction between TBCE and the microtubule-severing protein Spastin. Moreover, treatment of muscles with the microtubule-depolymerizing drug nocodazole implicated TBCE as a tubulin polymerizing protein. Taken together, these results demonstrate that TBCE is required for the normal development and function of ...

The contributions of MT attachment and kinetochore tension in the SAC are still under investigation (Nicklas et al., 1995; Waters et al., 1998; King et al., 2000; Shannon et al., 2002; Pinsky and Biggins, 2005), so the importance of tension was tested using brief treatment with taxol. This approach maintains MT attachment but reduces tension at the kinetochores of aligned chromosomes (Waters et al., 1998). Interkinetochore distance measurements confirmed decreased tension but not the completely relaxed kinetochore state observed in nocodazole (Fig. S4). PT89 signal was retained at the kinetochores of taxol-treated cells (Fig. 6 A) at levels similar to nocodazole-treated cells. As in nocodazole-treated cells, the retention of PT89 was linked to the retention of BubR1. Together, these results suggest that effective dynein dephosphorylation requires both MT attachment and kinetochore tension.. The requirement for both MT attachment and tension was reminiscent of previous work on kinetochore ...

Abstract Background Many cloned animals have been created by transfer of differentiated cells at G0/G1 or M phase of the cell cycle into enucleated M II oocytes having high maturation/meiosis/mitosis-promoting factor activity. Because maturation/meiosis/mitosis-promoting factor activity during oocyte maturation is maximal at both M I and M II, M I oocytes may reprogram differentiated cell nuclei as well. The present study was conducted to examine the developmental ability in vitro of porcine embryos reconstructed by transferring somatic cells (ear fibroblasts) into enucleated M I or M II oocytes. Results Analysis of the cell cycle stages revealed that 91.2 ± 0.2% of confluent cells were at the G0/G1 phase and 54.1 ± 4.4% of nocodazole-treated cells were at the G2/M phase, respectively. At 6 h after activation, nuclear swelling was observed in 50.0-88.9% and 34.4-39.5% of embryos reconstituted with confluent cells and nocodazole-treated cells regardless of the recipient oocytes, ...

Tyrosine phosphorylation is implicated in the formation, maintenance and turnover of cell-matrix adhesions (Barry and Critchley, 1994; Ridley and Hall, 1994; Chrzanowska-Wodnicka and Burridge, 1994; Bershadsky et al., 1996; Retta et al., 1996; Ayalon and Geiger, 1997; Schneider et al., 1998), yet the mechanism underlying this involvement is still obscure. In this study, we addressed this issue by comparing local changes in PY levels to the recruitment of several FA proteins induced by the microtubule-disrupting drug nocodazole. For this purpose, we have combined quantitative immunofluorescence microscopy with a novel approach for monitoring kinetics of tyrosine phosphorylation in live cells.. FA assembly is a multistage process that involves the transformation of small, dot-like focal complexes into large FAs (see Geiger et al., 2001). Previous studies have shown that application of mechanical force to focal complexes, either by increasing cytoskeletal contractility or by external perturbation, ...

Apoptosis resistance is the major cause of chemotherapy failure in most kinds of cancers, including Burkitt lymphomas (BL). To elucidate molecular mechanisms regulating the development of apoptosis resistance, a panel of 15 BL cell lines was investigated for apoptosis induction upon treatment with microtubule inhibitors taxol, nocodazole and vincristine. Significant differences were observed in the extent of apoptosis induction among BL cell lines examined. Interestingly, cell lines exhibiting resistance to taxol- or nocodazole-induced apoptosis, showed development of polyploidy (,4N) and vice versa, displaying an inverse relationship between apoptosis and polyploidy induction. Further, in sensitive cell lines taxol-induced apoptosis was accompanied by caspase activation, Bid cleavage and Mcl-1 down-regulation. In contrast, most apoptosis resistant cell lines exhibited a loss of Bax and Bak expression and showed prolonged mitotic arrest with ,4N DNA content upon treatment. To gain mechanistic ...

We hypothesized that the effects of Centrin2 inactivation could be associated with transient microtubule disruption. In support of this idea, we found that the treatment of freshly isolated E16 mouse neurons with the microtubule-destabilizing agent nocodazole (6 μm, 2 h) lead to disruption of the cytoplasmic polarity, as evidenced by the dispersion of the Golgi apparatus and late endosomes (see supplemental Fig. 3C,D, available at www.jneurosci.org as supplemental material) (n = 90 cells from 3 brains) as well as neurite retraction (data not shown) (de Anda et al., 2005). To confirm the specificity of the Centrin2 inactivation using CALI, we imaged neurons in irradiated slices that did not express the Centrin2-KR and found normal neuronal differentiation with the polarized cytoplasm oriented toward the medial/apical neurite (Fig. 3C,D) (8 h) that would become the axon (Fig. 3D, 42 h, black arrowheads and Inset 1) (for the whole time-lapse sequence and detailed z-stack analysis of the L1 ...

Initially, MTs in the vegetal shear zone are disorganized and short, but as cortical rotation progresses, they lengthen into parallel bundles, which have their plus-ends oriented away from the site of sperm entry (Fig. 2C,F) (Houliston and Elinson, 1991a). The subcortical MTs depolymerize near the end of the first cell cycle, ending the process of cortical rotation (Schroeder and Gard, 1992; Marrari et al., 2003). During rotation, the cortex (and the dorsalizing activity) moves towards the plus-ends of the MTs, explaining why the direction of cortical rotation depends on the site of fertilization and why the dorsal organizer forms on the side of the embryo opposite the sperm entry point (reviewed by Houliston and Elinson, 1992).. The proper formation of the MT array is crucial for the development of the dorsal axis. If MT polymerization is blocked during the first cell cycle by exposure to UV irradiation or to the MT-depolymerizing drug nocodazole, or with inhibitory antibodies to the ...

Stabilization of microtubules through the expression of Syk. (A)MDA-MB-231 cells lacking Syk (−Syk) or expressing Syk (+Syk)and treated with nocodazole were s

Spindle checkpoint proteins monitor the interaction of the spindle apparatus with the kinetochores, halting anaphase even if the microtubule attachment of only a single chromosome is altered. In this study, we show that Bub3p of Saccharomyces cerevisiae, an evolutionarily conserved spindle checkpoint protein, exhibits distinct interactions with an altered or defective kinetochore(s). We show for the first time that green fluorescent protein-tagged S. cerevisiae Bub3p (Bub3-GFP) exhibits not only a diffuse nuclear localization pattern but also forms distinct nuclear foci in unperturbed growing and G2/M-arrested cells. As Bub3-GFP foci overlap only a subset of kinetochores, we tested a model in which alterations or defects in kinetochore or spindle integrity lead to the distinct enrichment of Bub3p at these structures. In support of our model, kinetochore-associated Bub3-GFP is enriched upon activation of the spindle checkpoint due to nocodazole-induced spindle disassembly, overexpression of the

Here, we report that a centrosomal protein FOR20 [FOP (FGFR1 (fibroblast growth factor receptor 1) oncogene protein)-like protein of molecular mass of 20 kDa; also named as C16orf63, FLJ31153 or PHSECRG2] can regulate the assembly and stability of microtubules. Both FOR20 IgG antibody and GST (glutathione S-transferase)-tagged FOR20 could precipitate tubulin from the HeLa cell extract, indicating a possible interaction between FOR20 and tubulin. FOR20 was also detected in goat brain tissue extract and it cycled with microtubule-associated proteins. Furthermore, FOR20 bound to purified tubulin and inhibited the assembly of tubulin in vitro. The overexpression of FOR20 depolymerized interphase microtubules and the depletion of FOR20 prevented nocodazole-induced depolymerization of microtubules in HeLa cells. In addition, the depletion of FOR20 suppressed the dynamics of individual microtubules in live HeLa cells. FOR20-depleted MDA-MB-231 cells displayed zigzag motion and migrated at a slower rate ...

KCNH2 encodes the Kv11.1 α-subunit that underlies the rapidly activating delayed-rectifier K+ current in the heart. Loss-of-function KCNH2 mutations cause long QT syndrome type 2 (LQT2), and most LQT2-linked missense mutations inhibit the trafficking of Kv11.1 channel protein to the cell surface membrane. Several trafficking-deficient LQT2 mutations (e.g., G601S) generate Kv11.1 proteins that are sequestered in a microtubule-dependent quality control (QC) compartment in the transitional endoplasmic reticulum (ER). We tested the hypothesis that the QC mechanisms that regulate LQT2-linked Kv11.1 protein trafficking are mutation-specific. Confocal imaging analyses of HEK293 cells stably expressing the trafficking-deficient LQT2 mutation F805C showed that, unlike G601S-Kv11.1 protein, F805C-Kv11.1 protein was concentrated in several transitional ER subcompartments. The microtubule depolymerizing drug nocodazole differentially affected G601S- and F805C-Kv11.1 protein immunostaining. Nocodazole caused G601S

In addition to direct tumor cell cytotoxicity, chemotherapy can mediate tumor reduction through immune modulation of the tumor microenvironment to promote anti-tumor immunity. Mature dendritic cells (DCs) play key roles in priming robust immune responses in tumor-bearing hosts. Here, we screened a p …

Analysis and theory development in cognitive psychology and science education study remain mainly isolated. AP biology programs in high school and accomplished a score of 4 or 5 5 on the AP Biology examination. These college students participated as part of their normal classroom activities. The nonmajors group (= 68) was composed of students enrolled …Read More. ...

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Fluorescence images of control and Si-RNA transfected HeLa cells treated with nocodazole and allowed to recover. Mitotic cells were stained for alpha-...

Cytoskeleton inhibitors were used to study morphogenesis in the pathogenic and dimorphic fungus Candida albicans Nocodazole is a specific microtubule inhibitor and chloropropham (CIPC), at high concentrations, is an inhibitor of microtubules and microfilaments. Distribution of microtubules and microfilaments was studied by immunofluorescence techniques using anti-tubulin antibody with FITC-conjugated secondary antibody, and by staining with Rh-phalloidin. Nocodazole did not arrest apical cell elongation at a concentration (20 µg ml−1) that inhibited nuclear division and migration. Cytoplasmic and nuclear microtubules disappeared within 30 min in filamentous cells under these conditions. However, the Rh-phalloidin-stained actin granules which were localized in the tips of filamentous cells, and the microfilaments, were arranged normally at this concentration of nocodazole. Growth, and normal distribution of microtubules and microfilaments, were inhibited by a high concentration (200 µg ml−1) of

The IPL1 gene is required for high-fidelity chromosome segregation in the budding yeast Saccharomyces cerevisiae. Conditional ipl1ts mutants missegregate chromosomes severely at 37 degrees C. Here, we report that IPL1 encodes an essential putative protein kinase whose function is required during the later part of each cell cycle. At 26 degrees C, the permissive growth temperature, ipl1 mutant cells are defective in the recovery from a transient G2/M-phase arrest caused by the antimicrotubule drug nocodazole. In an effort to identify additional gene products that participate with the Ipl1 protein kinase in regulating chromosome segregation in yeast, a truncated version of the previously identified DIS2S1/GLC7 gene was isolated as a dosage-dependent suppressor of ipl1ts mutations. DIS2S1/GLC7 is predicted to encode a catalytic subunit (PP1C) of type 1 protein phosphatase. Overexpression of the full-length DIS2S1/GLC7 gene results in chromosome missegregation in wild-type cells and exacerbates the ...

We have examined the cortex of Caenorhabditis elegans eggs during pseudocleavage (PC), a period of the first cell cycle which is important for the generation of asymmetry at first cleavage (Strome, S. 1989. Int. Rev. Cytol. 114: 81-123). We have found that directed, actin dependent, cytoplasmic, and cortical flow occurs during this period coincident with a rearrangement of the cortical actin cytoskeleton (Strome, S. 1986. J. Cell Biol. 103: 2241-2252). The flow velocity (4-7 microns/min) is similar to previously determined particle movements driven by cortical actin flows in motile cells. We show that directed flows occur in one of the daughters of the first division that itself divides asymmetrically, but not in its sister that divides symmetrically. The cortical and cytoplasmic events of PC can be mimicked in other cells during cytokinesis by displacing the mitotic apparatus with the microtubule polymerization inhibitor nocodazole. In all cases, the polarity of the resulting cortical and ...

We constructed complexes between isolated chromosomes and microtubules made from purified tubulin to study the movement of chromosomes towards the minus end of microtubules in vitro, a process analogous to the movement of chromosomes towards the pole of the spindle at anaphase of mitosis. Our results show that the energy for this movement is derived solely from microtubule depolymerization, and indicate that anaphase movement of chromosomes is both powered and regulated by microtubule depolymerization at the kinetochore ...

Subsets of microtubules enriched in posttranslationally detyrosinated (Gundersen, G. G., M. H. Kalnoski, and J. C. Bulinski. 1984. Cell. 38:779) or acetylated (Piperno, G., M. Le Dizet, and X. Chang. 1987. J. Cell Biol. 104:298), alpha tubulin have previously been described in interphase cultured cells. In this study an immunofluorescence comparison of these minor populations of microtubules revealed that, in African green monkey kidney epithelial cells (TC-7 line), the population of microtubules enriched in detyrosinated tubulin was virtually coincident with the population enriched in acetylated alpha tubulin. In some cell types, however, such as human HeLa or marsupial PtK-2 cells, only one posttranslationally modified form of tubulin, i.e., acetylated or detyrosinated, respectively, was detectable in microtubules. In TC-7 cells, although both modifications were present, dissimilar patterns and kinetics of reappearance of microtubules enriched in detyrosinated and acetylated tubulin were ...

Although mitochondria are known to move actively in plant cells, little is known about how they move. In higher plants, actin filaments have been reported to be involved in the movement of organelles, such as chloroplasts, peroxisomes, endoplasmic reticulum and Golgi apparatus. Mitochondria were visualized in living ,i,Arabidopsis thaliana,/i, plants using fluorescent proteins fused to a mitochondria targeting signal. To compare the movement of mitochondria to the movements of another organelle, we also examined peroxisomes because of their similarity in size. Velocities of individual mitochondria and peroxisomes, as measured by time-lapse laser scanning microscopy, varied, although the average velocities of the two organelles were similar. Latrunculin B, an actin-depolymerizing drug, stopped movement of mitochondria and peroxisomes, demonstrating that movement of these organelles depends on actin filaments. On the other hand, propyzamide, a microtubule-depolymerizing drug, did not affect the ...

Similar to that described in previous studies (6, 7), our examination into cell cycle-dependent regulation of homologous recombination was undertaken by α-factor treatment to induce G1 arrest and nocodazole treatment to induce G2/M arrest. It has been reported that activation of homologous recombination is also observed in the S phase in S. cerevisiae (1, 20). Because replication stress is a major source of DSBs, DNA damage repair by homologous recombination is critically required to maintain genome stability in the S phase. Thus, it is reasonable to assume that a mechanism exists whereby homologous recombination is also activated in the S phase. In this study, we found that the phosphorylation of Rad51 and Rad52 for the progression of homologous recombination is catalyzed by Cdc28 combined with either Clb2 or Clb3. Given that Clb2 and Clb3 are redundantly expressed in the S and G2/M phases (fig. S2D), it is presumable that Cdc28 combined with either Clb2 or Clb3 induces Rad51 and Rad52 to ...

Based on these findings, we believe that microtubules are a critical target of PD environmental toxins such as rotenone, said Feng. Since many microtubule-depolymerizing agents are compounds naturally produced in many plants, our research points to the need to examine their possible link to Parkinsons disease. In addition, PD has a higher incidence in rural areas and is associated with pesticides and insecticides frequently used in farming practices ...

El nostre treball demostra que el 2ME2 inhibeix langiogènesis tumoral mitjançant la inhibició dels nivells proteics del factor HIF-1 així com la seva activitat transcripcional, incloent la transcripció del VEGF, en diverses línies cel.lulars. El 2ME2 no indueix la degradació proteica a través del proteosoma ni tampoc redueix els nivells dARNm del HIF-1, sinó que més aviat inhibeix la síntesi proteica de novo. Notablement, hem mostrat que el 2ME2 inhibeix el factor HIF-1 amb posterioritat a la disrupció del citoesquelet de microtúbuls aportant sòlides proves que afavoreixen una relació funcional entre els efectes anti-angiogènics i la disrupció dels microtúbuls. Aquests efectes no són una propietat única del 2ME2 sinó que és compartida per altres agents que actuen sobre els microtúbuls, com ara el taxol i la vincristina, suggerint un mecanisme dacció semblant per a tots aquests agents. Finalment, la nostra investigació demostra per primera vegada que el 2ME2 ...

Immunocompromisation is a major cause of opportunistic infections, activation of latent viruses, and the development of certain carcinomas such as Kaposis sarcoma (1, 2). This is particularly evident in many diseases that weaken the immune surveillance system, including AIDS and various cancer treatment modalities (e.g., radiation therapy, chemotherapy, and, in particular, antimicrotubule therapy), primarily because of severe myelosuppression and immunosuppression (3, 4). Finally, the resolution of these infections is dependent on the treatment modality of the malignancy and the ability of the host to mount an adequate immune response to fight off the infection (2).. Although antimicrotubule drugs constitute an important major class of anticancer drugs mainly typified by agents that either overpolymerize or bundle microtubules (such as the taxane family, ref. 5) or those that depolymerize them (such as the vincas; ref. 6, recently reviewed in refs. 7, 8), these agents, due to their gross ...

マウス・モノクローナル抗体 ab56676 交差種: Rat,Rb,Hu 適用: WB,IHC-P,Flow Cyt…Tubulin抗体一覧…画像、プロトコール、文献などWeb上の情報が満載のアブカムの Antibody 製品。国内在庫と品質保証制度も充実。

The incidence and mortality of breast cancer have increased throughout the world (Althuis et al., 2005), and it remains the most common cancer in women (Murray et al., 2012). In recent years, more and more drugs explored in clinical trials have provided promising advances in breast cancer treatments. Microtubule-binding agents are among the efficacious chemotherapeutic drugs that are commonly used for the treatment of breast cancer. Although antimicrotubule drugs, such as PTX and docetaxel, have been successfully applied in the clinic, drug resistance often occurred (Kavallaris, 2010) and acted as an obstacle for first-line chemotherapy. Augmented efflux transporters and diverse β-tubulin isotypes were involved in the development of drug resistance (Dumontet and Sikic, 1999). Thus, there is a pressing need to develop new microtubule inhibitors that can escape from the efflux of transporters related to MDR.. As a promising microtubule inhibitor candidate, DPT possesses various pharmacologic ...

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Lipid droplets (LDs) are intracellular organelles that provide fatty acids (FAs) to cellular processes including synthesis of membranes and production of metabolic energy. While known to move bidirectionally along microtubules (MTs), the role of LD motion and whether it facilitates interaction with other organelles are unclear. Here we show that during nutrient starvation, LDs and mitochondria relocate on detyrosinated MT from the cell centre to adopt a dispersed distribution. In the cell periphery, LD-mitochondria interactions increase and LDs efficiently supply FAs for mitochondrial beta-oxidation. This cellular adaptation requires the activation of the energy sensor AMPK, which in response to starvation simultaneously increases LD motion, reorganizes the network of detyrosinated MTs and activates mitochondria. In conclusion, we describe the existence of a specialized cellular network connecting the cellular energetic status and MT dynamics to coordinate the functioning of LDs and mitochondria ...

Biological activities of the 1,4-benzoquinone derivatives 5-O-ethylembelin (1) and 5-O-methylembelin (2) were investigated. Both of them showed anti proliferative activity against a panel of human tumor cell lines upon comparison to normal marsupial kidney cells (PtK2). They arrested HL-60 cells in the G(0)/G(1) phase of the cell cycle in a dose- and time-dependent manner. In HeLa cells, exposure to 100 mu M of 1 or 2 for 6 h induced a complete disassembly of the microtubule network and an increased number of cells blocked in mitotic stages. Treatment with 10 mu M of 1 and 2 for 24 h induced apoptosis in HL-60 cells. This evidence suggests that both 1 and 2 are promising novel antimitotic and anticancer molecules targeting microtubular proteins. ...

SID-530 is an intravenous formulation containing docetaxel, a semi-synthetic, second-generation taxane derived from a compound found in the European yew tree, Taxus baccata, with potential antineoplastic activity. Taxol analogue SID 530 binds to and stabilizes tubulin, inhibiting microtubule disassembly, which results in cell-cycle arrest at the G2/M phase and cell death. Check for active clinical trials or closed clinical trials using this agent. (NCI Thesaurus) (last updated: 6/23/2015)

SMCC-DM1 (DM1-SMCC) is a drug-linker conjugate composed of a potent microtubule-disrupting agent DM1 and a linker SMCC to make antibody drug conjugate (ADC). - Mechanism of Action & Protocol.

Motor proteins in the kinesin-8 family depolymerize microtubules in a length-dependent manner that may be crucial for controlling the length of organelles

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for 8 hours (by this time most MCF-7 cells had not yet accumulated in mitosis and thus remained adherent), fixed, stained with anti-α-tubulin, and visualized by fluorescence microscopy ...

Rudiger, M., Wehland, J., Weber, K. (1994). „The carboxy-terminal peptide of detyrosinated α tubulin provides a minimal system to study the substrate specificity of tubulin-tyrosine ligase. Eur. J. Biochem. 220: 309-320. PMID 7510228 ...

It is well known that c-Jun N-terminal kinase (JNK) plays pivotal roles in various mitotic events, but its function in mammalian oocyte meiosis remains unknown. In this study, we found that no specific JNK2 signal was detected in germinal vesicle stage. JNK2 was associated with the spindles especially the spindle poles and cytoplasmic microtubule organizing centers at prometaphase I, metaphase I, and metaphase II stages. JNK2 became diffusely distributed and associated with the midbody at telophase I stage. Injection of myc-tagged JNK2α1 mRNA into oocytes also revealed its localization on spindle poles. The association of JNK2 with spindle poles was further confirmed by colocalization with the centrosomal proteins, γ-tubulin and Plk1. Nocodazole treatment showed that JNK2 may interact with Plk1 to regulate the spindle assembly. Then we investigated the possible function of JNK2 by JNK2 antibody microinjection and JNK specific inhibitor SP600125 treatment. These two manipulations caused ...

Eribulin mesylate (eribulin), an analogue of the marine natural product halichondrin B, is a microtubule-depolymerizing drug that has utility in the treatment of patients with breast cancer. Clinical trial results have demonstrated that eribulin treatment provides a survival advantage to patients with metastatic or locally advanced breast cancer previously treated with an anthracycline and a taxane. Furthermore, a pooled analysis of two pivotal phase III trials has demonstrated that eribulin also improves overall survival in several patient subgroups, including in women with HER2-negative disease and triple-negative breast cancer. This review covers the preclinical research that led to the clinical testing and approval of eribulin, as well as subsequent research that was prompted by distinct and unexpected effects of eribulin in the clinic. Initial studies with halichondrin B demonstrated unique effects on tubulin binding that resulted in distinct microtubule-dependent events and antitumor ...

The trafficking of the adenomatous polyposis coli (APC) tumour suppressor protein in mammalian cells is a perennially controversial topic. Immunostaining evidence for an actin-associated APC localisation at intercellular junctions has been previously presented, though live imaging of mammalian junctional APC has not been documented. Using live imaging of transfected COS-7 cells we observed intercellular junction-associated pools of GFP-APC in addition to previously documented microtubule-associated GFP-APC and a variety of minor localisations. Although both microtubule and junction-associated populations could co-exist within individual cells, they differed in their subcellular location, dynamic behaviour and sensitivity to cytoskeletal poisons. GFP-APC deletion mutant analysis indicated that a protein truncated immediately after the APC armadillo repeat domain retained the ability to localise to adhesive membranes in transfected cells. Supporting this, we also observed junctional APC immunostaining in

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To determine whether TRF function was required to maintain cohesion, we examined cohesion in the trf4-ts trf5double mutant. If TRF function were required to maintain cohesion, then cells arrested with nocodazole at a permissive temperature and then shifted to the nonpermissive temperature should show an increased cohesion defect. After nocodazole arrest at the permissive temperature of 30°C, the trf4-ts trf5double mutant was able to maintain cohesion as monitored by the GFP-marked chromosome assay. However, when mutant cells were shifted to 37°C for 3 hours, a marked increase in cohesion-defective cells was observed (Fig. 3B). This demonstrates that TRF function is indeed required to maintain cohesion.. The ability of cells lacking TRF function to duplicate the genome was then examined as follows. Synchronous populations of either wild-type or otherwise isogenic trf4-ts trf5double-mutant G1 daughter cells were obtained by centrifugal elutriation. Wild-type and mutant cells were then released ...

4SC-207 is a novel microtubule inhibitor, which shows strong anti-proliferative activity in a large panel of tumor cell lines with an average GI50 of 11 nM. In particular, 4SC-207 is active in multi-drug resistant cell lines, such as HCT-15 and ACHN, suggesting that it is a poor substrate for drug efflux pumps. 4SC-207 inhibits microtubule growth in vitro and in vivo and promotes, in a dose dependent manner, a mitotic delay/arrest, followed by apoptosis or aberrant divisions due to chromosome alignment defects and formation of multi-polar spindles. Furthermore, preliminary data from preclinical studies suggest low propensity towards bone marrow toxicities at concentrations that inhibit tumor growth in paclitaxel-resistant xenograft models. 4SC-207 may be a potential anti-cancer agent.

TY - JOUR. T1 - Organization of non-centrosomal microtubules in epithelial cells. AU - Toya, Mika. AU - Takeichi, Masatoshi. PY - 2016/1/1. Y1 - 2016/1/1. N2 - Polarized epithelial cells contain a characteristic array of microtubules in which non-centrosomal microtubules are aligned along the apical-to-basal axis of the cell with their minus ends oriented towards the apical pole. Although this unique orientation of microtubules was discovered in the late 1980s, how this orientation is established remains unresolved partly because of limited information about molecular factors that regulate the minus ends of non-centrosomal microtubules. Recent studies, however, identified novel minus end- associated proteins, revealing mechanisms by which the polarized arrays of microtubules are established in epithelial cells. These studies have also demonstrated the importance of apico-basally orientated microtubules in intra-structural organization of cells. This review focuses on recent progress of our ...

The cytoskeleton is responsible for cell shape, motility (movement) of the cell as a whole, and motility of organelles within a cell. There are three types of filaments in the cytoplasm of most vertebrate cells: microfilaments, microtubules, and intermediate filaments.. filament systems are able to lengthen or shorten very rapidly. This dynamic nature of the cytoskeleton is necessary for cells to be able to change shape, complete cell division, or migrate.. The microfilament system is a network of filaments 6 nanometers (nm) in diameter that are important for anchoring plasma membrane proteins, for producing cell movement, and for cell division. The base filament is composed of a protein called actin.. Microtubules are the largest of the cytoskeletal filaments with a diameter of 25 nm. There are many parallels between the microfilament cytoskeletal system and the microtubule system. Like microfilaments, microtubules are produced by the self-assembly of a subunit, which in the case of ...

MLN0264 is an investigational antibody-drug conjugate (ADC) that consists of the human anti-guanylyl cyclase C (GCC) antibody linked to a microtubule-disrupting agent (monomethyl auristatin). As ADCs have a very long clearance half-life, the potential exists for a highly infrequent dosing schedule. A quantitative understanding of the relationship between exposure and preclinical antitumor biological activity is thus applied to support dose schedule selection in the clinic.. In this study, we develop a pharmacokinetic/efficacy (PK/E) relationship in xenograft models to evaluate the predictive contributions of exposure and xenograft characteristics to MLN0264 biological activity. Single dose pharmacokinetic (PK) data were obtained for a range of time points, and a linear two-compartment PK model was built. Xenograft biological activity studies were conducted in which MLN0264 was administered at various dose levels and dosing schedules to mice bearing one of six different xenograft models. We used ...

Mitotic spindle position defines the cell cleavage site during cytokinesis. cortex. In elongating cells asymmetrically, dynein-dependent spindle anchoring at the fixed cell cortex guarantees correct spindle setting. Our outcomes reveal the anaphase-specific spindle centering systems that obtain equal-sized cell department. syncytial embryos (Silverman-Gavrila et al., 2008). To evaluate the contribution of chromosome-derived Ran-GTP indicators in mammalian cells, we utilized tsBN2 cells, which include a heat range delicate mutation in RCC1 that stops the formation of Ran-GTP at the restricted heat range (Nishitani et al., 1991). In nocodazole treated tsBN2 cells, Anillin was decreased from the cell cortex in the location of the chromosome herd at the permissive heat range (Fig. 7A, still left) equivalent to HeLa cells (Fig. 6E). Nevertheless, at the restricted heat range, Anillin localised to the cell cortex also in the location of chromosomes (Fig. 7A, correct). The heat range change do not ...

Mitotic spindle position defines the cell cleavage site during cytokinesis. cortex. In elongating cells asymmetrically, dynein-dependent spindle anchoring at the fixed cell cortex guarantees correct spindle setting. Our outcomes reveal the anaphase-specific spindle centering systems that obtain equal-sized cell department. syncytial embryos (Silverman-Gavrila et al., 2008). To evaluate the contribution of chromosome-derived Ran-GTP indicators in mammalian cells, we utilized tsBN2 cells, which include a heat range delicate mutation in RCC1 that stops the formation of Ran-GTP at the restricted heat range (Nishitani et al., 1991). In nocodazole treated tsBN2 cells, Anillin was decreased from the cell cortex in the location of the chromosome herd at the permissive heat range (Fig. 7A, still left) equivalent to HeLa cells (Fig. 6E). Nevertheless, at the restricted heat range, Anillin localised to the cell cortex also in the location of chromosomes (Fig. 7A, correct). The heat range change do not ...

Subcellular distribution of mass can be analyzed by a technique that involves culturing cells on interferometers and digitizing their interference contours. Contour sampling resulted in 102 variables per cell, which were predictors of oncogenic transformation. Cell phenotypes can be deconstructed by use of latent factors, which represent the covariance of the real variables. The reversal of the cancertype phenotype by a combination of microtubule- stabilizing and -depolymerizing agents was described previously. The implications of these results have been explored by clinicians who treated patients with the combination of docetaxel and vinorelbine (Navelbine®). The current study was performed to determine the effects of different combinations on phenotype and in phases of the cell cycle other than mitosis. Combinations of paclitaxel with either colchicine, podophyllotoxin, nocodazole, or vinblastine caused phenotype reversal. Paclitaxel analogue, 7-deoxytaxol, by itself caused reversal. Factors #4,

The table lists cell cycle regulated chromatin remodeling genes. Five transcriptome data sets, in which the genes that are periodically expressed in at least one data set, and one ribosomal profiling (translationally regulated) data set are included. Periodically expressed genes are indicated with cell cycle phase in which they show the highest expression level. Gene sets in which a given gene is not significantly cell cycle regulated is indicated as NR (not transcriptionally regulated in the cell cycle) or NTR (not translationally regulated in the cell cycle) in transcriptome and ribosomal profiling data sets, respectively. Four experiments are supplemented with expression profiles which can be found by clicking the gene name. Synchronization methods are abbreviated as: SS=serum starvation, TT= double thymidine block, NZ=nocodazole, MS=mitotic shake-off ...