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1

Cellular organelle transport and positioning by plasma streaming

100%
Wanka F., Van Zoelen E. J. J.
Cellular and Molecular Biology Letters
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2003
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tom 08
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nr 4
Our analysis of known data reveals that translocations of passively movable cellular organelles from tiny granules up to large cell nuclei can be ascribed to transport by streaming cytoplasm. The various behaviours, such as velocity changes during more or less interrupted movements, forth and back shuttling and particle rotation result from different types of plasma circulation. Fast movements over long distances, as observed in the large characean internodial cells occur in strong streams generated by myosin in bundles of actin filaments in the direction of the barbed filament ends. Slow movements with frequent reversions of the direction are typical for neuronal axons, in which an anterograde plasma flow, produced in a thin layer of membrane-attached actin filaments, is compensated by a retrograde stream, produced by dynein activity in the central bundle of microtubules. Here particle rotation is due to steep flow velocity gradients, and frequent changes of particle movements result from minor particle displacements in radial directions. Similar shuttling of pigment granules in the lobes of epidermal chromatophores results from the same mechanism, whereby the centrifugal movement along astral microtubules is due to flow generated by excess of kinesin activity and the centripetal movement to the plasma recycling through the intermicrotubular space. If the streaming pattern is reversed by switching to excess dynein activity, the moving granules are trapped in the high microtubule density at the aster center. The presence of larger bodies in asters disturbs the regular, kinesin-dependent microtubule distribution in such a way that a superimposed centrifugal plasma flow develops in the microtubule-dense layer along them, which is recycled in the microtubule-free space, created by their presence. Consequently, at excess kinesin activity, nuclei, mitochondria as well as chromosome fragments move towards the aster center until they reach a dynamically stabilized position that depends on the local microtubule density. These various behaviours are not rationally explainable by models based on a mechanical stepping along microtubules or actin filaments.
2
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Stabilization of actin filaments with phalloidin has no influence on chloroplast movement in Arabidopsis mesophyll cells

84%
Krzeszowiec W., Jagiello-Flasinska D., Gabrys H.
BioTechnologia. Journal of Biotechnology Computational Biology and Bionanotechnology
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2013
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tom 94
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nr 3
3

Establishing and functional characterization of an HEK-293 cell line expressing autofluorescently tagged beta-actin [pEYFP-actin] and the neurokinin type 1 receptor [NK1-R]

84%
Hrovat A., Zavec A. B., Pogacnik A., Frangez R., Vrecl M.
Cellular and Molecular Biology Letters
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2010
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tom 15
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nr 1
55-69
This study focused on establishing and making a comprehensive functional characterization of an HEK-293-transfected cell line that would coexpress the enhanced yellow fluorescent protein-actin (pEYFP-actin) construct and the neurokinin type 1 receptor (NK1-R), which is a member of the seven transmembrane (7TM) receptor family. In the initial selection procedure, the cloning ring technique was used alone, but failed to yield clones with homogenous pEYFP-actin expression. Flow cytometry sorting (FCS) was subsequently used to enrich the pEYFP-actin-expressing subpopulation of cells. The enzyme-linked immunosorbent assay (ELISA), FCS and quantitative real-time reverse transcription/polymerase chain reaction (RT-PCR) were then employed to monitor the passage-dependent effects on transgene expression and to estimate the total β-actin/pEYFP-actin ratio. NK1-R was characterized via radioactive ligand binding and the second messenger assay. The suitability of the pEYFP-actin as a marker of endogenous actin was assessed by colocalizing pEYFP-actin with rhodamine-phalloidine-stained F-actin and by comparing receptor- and jasplakinolide-induced changes in the actin cytoskeleton organization. These experiments demonstrated that: i) both constructs expressed in the generated transfected cell line are functional; ii) the estimated pEYFP-actin: endogenous β-actin ratio is within the limits required for the functional integrity of the actin filaments; and iii) pEYFP-actin and rhodamine-phalloidine-stained F-actin structures colocalize and display comparable reorganization patterns in pharmacologically challenged cells.
4

The role of actin and microtubule networks in plasmid DNA intracellular trafficking

84%
Ondrej V., Likasova E., Falk M., Kozubek S.
Acta Biochimica Polonica
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2007
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tom 54
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nr 3
This work is focused on the function of the microtubule and actin networks in plasmid DNA transport during liposomal transfection. We observed strong binding of plasmid DNA-lipid complexes (lipoplexes) to both networks and directional long-range motion of these lipoplexes along the microtubules. Disruption of either of these networks led to the cessation of plasmid transport to the nucleus, a decreased mobility of plasmids, and accumulation of plasmid DNA in large aggregates at the cell periphery. Our findings show an indispensable but different role of both types of cytoskeleton, actin and microtubular, in the processes of gene delivery.
5

Force generation by cellular motors

84%
Wanka F., Van Zoelen E. J. J.
Cellular and Molecular Biology Letters
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2003
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tom 08
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nr 4
Cell motility processes in non-muscle cells depend on the activity of motor proteins that bind to either microtubules or actin filaments. From presently available data it must be concluded that the driving force is generated by transient interaction of the respective motors with microtubules or actin filaments which then activates the binding and hydrolysis of ATP. This reaction results in an abrupt discharge of the motor molecule, the direction of which is determined by the spatial orientation of its binding to the helical and polar vehicle. The latter is thereby propelled in its length direction and simultaneously undergoes an axial rotation, while the expelled motor exerts an oppositely directed current in the surrounding fluid, comparable to jet propulsion. Force production, propulsion velocities and energy requirements known from in vitro studies comply with those derived from the theory. The theory opens new ways for the understanding of cellular activities such as particle transport, mitosis and morphodynamics.
6

Motility assay: achievements and perspectives

84%
Makuch R., Stepkowski D.
Acta Biochimica Polonica
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1993
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tom 40
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nr 3
7

Visualisation of microtubules and actin filaments in fixed BY-2 suspension cell

67%
Szechynska-Hebda M., Wedzony M., Dubas E., Emons A. M., Lammeren A.
Biological Letters
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2005
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tom 42
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nr 2 Spec.Vol.
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