ALBERT — All Library Books, journals and Electronic Records Telegrafenberg

1

Electronic Resource

Characterization of tetramethylrhodaminyl-phalloidin binding to cellular F-actin (1992)

Cano, Manuel L. ; Cassimeris, Lynne ; Joyce, Michael ; [et al.]

New York, NY : Wiley-Blackwell

Cell Motility and the Cytoskeleton 21 (1992), S. 147-158

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ISSN: 0886-1544

Keywords: depolymerization ; DNase I ; association rate constant ; dissociation rate constant ; polymor-phonuclear leukocyte ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Medicine

Notes: Fluorescent derivatives of phallcidin are widely used to measure filamentous actin (F-actin) levels and to stabilize F-actin. We have characterized the kinetics and affinity of binding of tetramethylrhodaminy (TRITC)-phalloidin to rabbit skeletal muscle F-actin and to F-actin in lysates of rabbit polymorphonuclear leukocytes (PMNs). We have defined conditions where TRITC-phalloidin can be used to inhibit F-actin depolymerization and to quantify F-actin without prior fixation. By equi librium measurements, the affinity of TRITC-phalloidin binding to rabbit skeletal muscle F-actin (pyrene labeled) or to PMN lysate F-actin was 1-4 × 10-7 M. In both cases, the stoichiometry of binding was approximately 1:1. Kinetic measurements of TRITC-phalloidin binding to PMN lysate F-actin resulted in an association rate constant of 420 ± 120 M-1 sec-1 and a dissociation rate constant of 8.3 ± 0.9 ± 10-5 sec-1. The affinity calculated from the kinetic measurements. (2 ± 1 × 10-7 M) agreed well with that obtained by equilibrium measurements. The rate with which 0.6 μM TRITC-phalloidin inhibited 0.1 μM pyrenyl F-actin depolymerization (90% inhibition in 10 sec) was much faster than the rate of binding to pyrenyl F-actin (〈1% bound in 10 sec), suggesting that phalloidin binds to filament ends more rapidly than to the rest of the filament. We show that TRITC-phalloidin can be used to measure F-actin levels in cell lysates when G-actin is also present (i.e., in cell lysates at high concentrations) if DNase I is included to prevent phalloidin-induced polymerization.

Additional Material: 9 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/cm.970210208

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2

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High hydrostatic pressure effects in vivo: Changes in cell morphology, microtubule assembly, and actin organization (1988)

Bourns, Brenda ; Franklin, Samuel ; Cassimeris, Lynne ; [et al.]

New York, NY : Wiley-Blackwell

Cell Motility and the Cytoskeleton 10 (1988), S. 380-390

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ISSN: 0886-1544

Keywords: stress fiber ; cytoskeleton ; microvilli ; tubulin ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Medicine

Notes: We present the first study of the changes in the assembly and organization of actin filaments and microtubules that occur in epithelial cells subjected to the hydrostatic pressures of the deep sea. Interphase BSC-1 epithelial cells were pressurized at physiological temperature and fixed while under pressure. Changes in cell morphology and cytoskeletal organization were followed over a range of pressures from 1 to 610 atm. At atmospheric pressure, cells were flat and well attached. Exposure of cells to pressures of 290 atm or greater caused cell rounding and retraction from the substrate. This response became more pronounced with increased pressure, but the degree of response varied within the cell population in the pressure range of 290-400 atm, Microtubule assembly was not noticeably affected by pressures up to 290 atm, but by 320 atm, few microtubules remained. Most actin stress fibers completely disappeared by 290 atm. High pressure did not simply induce the overall depolymerization of actin filaments for, concurrent with cell rounding, the number of visible microvilli present on the cell surface increased dramatically. These effects of high pressure were reversible. Cells re-established their typical morphology, microtubule arrays appeared normal, and stress fibers reformed after approximately 1 hour at atmospheric pressure. High pressure may disrupt the normal assembly of microtubules and actin filaments by affecting the cellular regulatory mechanisms that control cytological changes during the transition from interphase into mitosis.

Additional Material: 7 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/cm.970100305

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3

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Chromosome fiber dynamics and congression oscillations in metaphase PtK2 Cells at 23°C (1991)

Wise, Dwayne ; Cassimeris, Lynne ; Rieder, Conly L. ; [et al.]

New York, NY : Wiley-Blackwell

Cell Motility and the Cytoskeleton 18 (1991), S. 131-142

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ISSN: 0886-1544

Keywords: mitosis ; microtubules ; tubulin incorporation ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Medicine

Notes: A bioriented chromosome is tethered to opposite spindle poles during congression by bundles of kinetochore microtubules (kMts). At room temperature, kinetochore fibers are a dominant component of mitotic spindles of PtK2 cells. PtK2 cells at room temperature were injected with purified tubulin covalently bound to DTAF and congression movements of individual chromosomes were recorded in time lapse. Congression movements of bioriented chromosomes between the poles occur over distances of 4.5 μm or greater. DTAF-tubulin injection had no effect on either the velocity or extent of these movements. Other cells were lysed, fixed, and the location of DTAF-tubulin incorporation was detected from digitally processed images of indirect immunofluorescence of an antibody to DTAF. Microtubules were labeled with an anti-beta tubulin antibody. At 2-5 minutes after injection, concentrated DTAF-tubulin staining was seen in the kinetochore fibers proximal to the kinetochores; a low concentration of DTAF-tubulin staining occurred at various sites through the remaining length of the fibers toward the pole. Kinetochore fibers in the same cell displayed different lengths (0.2 to 4 μm) of concentrated DTAF-tubulin incorporation proximal to the kinetochore, as did sister kinetochore fibers. Ten minutes after injection, the lengths of DTAF-containing chromosomal fibers were greater than expected if incorporation resulted solely from the lengthening of kinetochore microtubules due to congression movements of the chromosomes. Besides incorporation as a result of chromosome movement, two other mechanisms might explain the length of the DTAF-containing segments: (1) a poleward flux of tubulin subunits (Mitchison, 1989) or (2) capture of DTAF-containing nonkinetochore microtubules.

Additional Material: 8 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/cm.970180208

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4

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Regulation of microtubule dynamic instability (1993)

Cassimeris, Lynne

New York, NY : Wiley-Blackwell

Cell Motility and the Cytoskeleton 26 (1993), S. 275-281

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ISSN: 0886-1544

Keywords: Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Biology , Medicine

Additional Material: 2 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/cm.970260402

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5

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Spectral analysis of microtubule assembly dynamics (1996)

Odde, David J. ; Buettner, Helen M. ; Cassimeris, Lynne

Hoboken, NJ : Wiley-Blackwell

AIChE Journal 42 (1996), S. 1434-1442

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ISSN: 0001-1541

Keywords: Chemistry ; Chemical Engineering

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Chemistry and Pharmacology , Process Engineering, Biotechnology, Nutrition Technology

Notes: Microtubules are linear polymers of the cytoskeleton that serve to organize the cytoplasm of eukaryotic cells. Understanding how microtubule polymers self-assemble is important in biotechnology, including the development of novel cancer therapies and proper guidance of regenerating neurons. The assembly of microtubules occurs by a unique process whereby an individual microtubule undergoes abrupt and apparently stochastic switching between alternating steady states of growth and shrinkage, a phenomenon known as microtubule dynamic instability. To characterize these oscillations spectral (frequency-domain) analysis, commonly used in engineering for system identification, was applied. Power spectra of the individual microtubule-length life histories revealed oscillations within growth phases, directly reflecting acceleration and deceleration in the growth process. These fluctuations were not accounted for by the standard two-state model commonly used in the analysis of microtubule assembly, despite the inclusion of simulated measurement error in the model. Thus, the spectral analysis of microtubule assembly permitted characterization of assembly process dynamics independent of particular assembly models, and as such represents a powerful analytical framework within which to study microtubule dynamic instability and assess its function in vivo.

Additional Material: 8 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/aic.690420524

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6

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Metal evaporation shadowing: A computer simulation (1985)

Colquhoun, William ; Sokol, Roger ; Davison, Edwin ; [et al.]

New York, NY : Wiley-Blackwell

Journal of Electron Microscopy Technique 2 (1985), S. 353-370

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ISSN: 0741-0581

Keywords: Metal evaporation ; Shadowing ; Computer simulation ; Resolution ; Grain ; Void space ; Life and Medical Sciences ; Cell & Developmental Biology

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Natural Sciences in General

Notes: This paper addresses the question of what is the very best deposition that can be achieved by metal evaporation shadowing. It deals, therefore, with the resolution attainable in metal shadowed specimens. The analysis disregards interatomic attractions (nucleation, crystallite formation, decoration) and generates depositions solely on the basis of probabilities of vacant, single, and multiple atom landings at different points on a substrate.The treatment starts by showing that random number generation can be used to simulate reasonably the evaporation of metal atoms from a shadowing source and their deposit on a substrate. A Fortran program tabulates and displays these simulated atom landings. The analysis shows that one can never expect to obtain a perfectly uniform deposit; some areas will always be heavily filled while others will be lightly filled or vacant. Resolution is thereby limited. The analysis shows how a deposit becomes more uniform as the number of atoms deposited increases. It predicts optimal amounts of deposition for various shadowing angles. The analysis also provides a rationale for the way in which deposition grain coarsens and resolution worsens as the angle of shadow is made more steep.Computer-generated “ideal” depositions are compared with real shadow grains achieved under equivalent experimental conditions. The simulations are also compared with deposits achieved by novel shadowing methods. Ways to improved metal evaporation technique are suggested by the analysis and these are discussed.

Additional Material: 12 Ill.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1002/jemt.1060020405

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