scholarly journals The CENP-S complex is essential for the stable assembly of outer kinetochore structure

2009 ◽  
Vol 186 (2) ◽  
pp. 173-182 ◽  
Author(s):  
Miho Amano ◽  
Aussie Suzuki ◽  
Tetsuya Hori ◽  
Chelsea Backer ◽  
Katsuya Okawa ◽  
...  
Keyword(s):  
Hela Cells ◽  
Protein S ◽  
Cell Analysis ◽  
Molecular Architecture ◽  
Outer Plate ◽  
Kinetochore Assembly ◽  
Full Complement ◽  
Dt40 Cells ◽  
Live Cell Analysis ◽  
Mitotic Behavior

The constitutive centromere-associated network (CCAN) proteins are central to kinetochore assembly. To define the molecular architecture of this critical kinetochore network, we sought to determine the full complement of CCAN components and to define their relationships. This work identified a centromere protein S (CENP-S)–containing subcomplex that includes the new constitutive kinetochore protein CENP-X. Both CENP-S– and CENP-X–deficient chicken DT40 cells are viable but show abnormal mitotic behavior based on live cell analysis. Human HeLa cells depleted for CENP-X also showed mitotic errors. The kinetochore localization of CENP-S and -X is abolished in CENP-T– or CENP-K–deficient cells, but reciprocal experiments using CENP-S–deficient cells did not reveal defects in the localization of CCAN components. However, CENP-S– and CENP-X–deficient cells show a significant reduction in the size of the kinetochore outer plate. In addition, we found that intrakinetochore distance was increased in CENP-S– and CENP-X–deficient cells. These results suggest that the CENP-S complex is essential for the stable assembly of the outer kinetochore.

Projected Structure of the Surface Protein of Sulfolobus Spec. B12 Determined to a Resolution of 1.0 NM by Cryo-Electronmicroscopy

1990 ◽  
Vol 48 (1) ◽  
pp. 102-103
Author(s):  
G. Lembcke ◽  
F. Zemlin
Keyword(s):  
Low Dose ◽  
Maximum Dose ◽  
Three Dimensional ◽  
Surface Protein ◽  
Protein S ◽  
Radiation Sensitivity ◽  
Specimen Preparation ◽  
Molecular Architecture ◽  
High Radiation ◽  
Fritz Haber

The thermoacidophilic archaebacterium Sulfolobus spec. B12 , which is closely related to Sulfolobus solfataricus , possesses a regularly arrayed surface protein (S-layer), which is linked to the plasma membrane via spacer elements spanning a distinct interspace of approximately 18 nm. The S-layer has p3-Symmetry and a lattice constant of 21 nm; three-dimensional reconstructions of negatively stained fragments yield a layer thickness of approximately 6-7 nm.For analysing the molecular architecture of Sulfolobus surface protein in greater detail we use aurothioglucose(ATG)-embedding for specimen preparation. Like glucose, ATG, is supposed to mimic the effect of water, but has the advantage of being less volatile. ATG has advantages over glucose when working with specimens composed exclusively of protein because of its higher density of 2.92 g cm-3. Because of its high radiation sensitivity electromicrographs has to be recorded under strict low-dose conditions. We have recorded electromicrographs with a liquid helium-cooled superconducting electron microscope (the socalled SULEIKA at the Fritz-Haber-lnstitut) with a specimen temperature of 4.5 K and with a maximum dose of 2000 e nm-2 avoiding any pre-irradiation of the specimen.

Wee1 Inhibition Enhances the Anti-Tumor Effects of Capecitabine in Preclinical Models of Triple-Negative Breast Cancer.

2020 ◽  
Author(s):  
Todd Pitts ◽  
Dennis M Simmons ◽  
Stacey M Bagby ◽  
Sarah J Hartman ◽  
Betelehem W Yacob ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Triple Negative Breast Cancer ◽  
Combination Treatment ◽  
Triple Negative ◽  
Single Agent ◽  
Cell Analysis ◽  
Preclinical Models ◽  
Pdx Models ◽  
Live Cell Analysis

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive subtype defined by lack of hormone receptor expression and non-amplified HER2. Adavosertib (AZD1775) is a potent, small molecule, ATP-competitive inhibitor of the Wee1 kinase that potentiates the activity of many DNA-damaging chemotherapeutics and is currently in clinical development for multiple indications. The purpose of this study was to investigate the combination of AZD1775 and capecitabine/5-FU in preclinical TNBC models. Methods: TNBC cell lines were treated with AZD1775 and 5-FU and cellular proliferation was assessed in real-time using IncuCyte® Live Cell Analysis. Apoptosis was assessed via the Caspase-Glo 3/7 assay system. Western blotting was used to assess changes in expression of downstream effectors. TNBC PDX models were treated with AZD1775, capecitabine, or the combination and assessed for tumor growth inhibition. Results: From the initial PDX screen, two of the four TNBC PDX models demonstrated a better response in the combination treatment than either of the single agents. As confirmation, two PDX models were expanded for statistical comparison . Both PDX models demonstrated a significant growth inhibition in the combination versus either of the single agents. (TNBC012, p<0.05 combo vs adavosertib or capecitabine, TNBC013, p<0.01 combo vs adavosertib or capecitabine ). An enhanced antiproliferative effect was observed in the adavosertib/5-FU combination treatment as measured by live cell analysis. An increase in apoptosis was observed in two of the four cell lines in the combination when compared to single agent treatment. Treatment with single agent adavosertib resulted in an increase in p-CDC2 in a dose dependent manner that was also observed in the combination treatment. Similar results were observed with γH2AX in two of the four cell lines tested. No significant changes were observed in Bcl-xL following treatment in any of the cell lines. Conclusions: The combination of adavosertib and capecitabine/5-FU demonstrated enhanced combination effects both in vitro and in vivo in preclinical models of TNBC. These results support the clinical investigation of this combination in patients with TNBC, including those with brain metastasis given the CNS penetration of both agents.

DNA Hydrogel with Aptamer-Toehold-Based Recognition, Cloaking, and Decloaking of Circulating Tumor Cells for Live Cell Analysis

Nano Letters ◽  
2017 ◽  
Vol 17 (9) ◽  
pp. 5193-5198 ◽  
Author(s):  
Ping Song ◽  
Dekai Ye ◽  
Xiaolei Zuo ◽  
Jiang Li ◽  
Jianbang Wang ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
Live Cell ◽  
Cell Analysis ◽  
Dna Hydrogel ◽  
Live Cell Analysis

Abstract 2156: Quantifying cell subsets and heterogeneity in living cultures using real time live-cell analysis

2019 ◽  
Author(s):  
Nicola Bevan ◽  
Tim Jackson ◽  
Clare Szybut ◽  
Lauren Kelsey ◽  
Hinnah Campwala ◽  
...  
Keyword(s):  
Real Time ◽  
Live Cell ◽  
Cell Analysis ◽  
Live Cell Analysis

Abstract 5266: Kinetic measurements of intracellular ATP levels in co-culture models using live-cell analysis

2019 ◽  
Author(s):  
Cicely L. Schramm ◽  
Michael L. Bowe ◽  
Laura A. Skerlos ◽  
Grigory S. Filonov ◽  
Yong X. Chen ◽  
...  
Keyword(s):  
Live Cell ◽  
Cell Analysis ◽  
Kinetic Measurements ◽  
Intracellular Atp ◽  
Atp Levels ◽  
Culture Models ◽  
Live Cell Analysis

Abstract 2156: Quantifying cell subsets and heterogeneity in living cultures using real time live-cell analysis

2019 ◽  
Author(s):  
Nicola Bevan ◽  
Tim Jackson ◽  
Clare Szybut ◽  
Lauren Kelsey ◽  
Hinnah Campwala ◽  
...  
Keyword(s):  
Real Time ◽  
Live Cell ◽  
Cell Analysis ◽  
Live Cell Analysis

Abstract 5266: Kinetic measurements of intracellular ATP levels in co-culture models using live-cell analysis

2019 ◽  
Author(s):  
Cicely L. Schramm ◽  
Michael L. Bowe ◽  
Laura A. Skerlos ◽  
Grigory S. Filonov ◽  
Yong X. Chen ◽  
...  
Keyword(s):  
Live Cell ◽  
Cell Analysis ◽  
Kinetic Measurements ◽  
Intracellular Atp ◽  
Atp Levels ◽  
Culture Models ◽  
Live Cell Analysis
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