scholarly journals Reactivation of organelle movements along the cytoskeletal framework of a giant freshwater ameba.

1986 ◽  
Vol 103 (2) ◽  
pp. 605-612 ◽  
Author(s):  
M P Koonce ◽  
M Schliwa
Keyword(s):  
Intracellular Transport ◽  
Cytoplasmic Dynein ◽  
Model System ◽  
Organelle Transport ◽  
Nucleoside Triphosphates ◽  
High Concentrations

The peripheral feeding network of the giant freshwater ameba Reticulomyxa can be easily and rapidly lysed to produce an extensive, stable, and completely exposed cytoskeletal framework of colinear microtubules and microfilaments. Most of the organelles that remain attached to this framework resume rapid saltatory movements at rates of up to 20 micron/s if ATP is added. This lysed model system is also capable of other forms of motility, namely an active splaying of microtubule bundles and bulk streaming. Reactivation does not occur with other nucleoside triphosphates, requires Mg ions, is insensitive to even high concentrations of erythro-9-(3-[2-hydroxynonyl]) adenine, is sensitive to vanadate only at concentrations of approximately 100 microM, and is inhibited by N-ethylmaleimide at concentrations greater than 100 microM. The physiology of this reactivation suggests an organelle transport motor distinct from cytoplasmic dynein and possibly the recently described kinesin. This system can serve as a model for elucidating the mechanisms of intracellular transport and, in addition, provides a unique opportunity to examine associations between microtubules and microfilaments.

scholarly journals Dynactin is required for bidirectional organelle transport

2003 ◽  
Vol 160 (3) ◽  
pp. 297-301 ◽  
Author(s):  
Sean W. Deacon ◽  
Anna S. Serpinskaya ◽  
Patricia S. Vaughan ◽  
Monica Lopez Fanarraga ◽  
Isabelle Vernos ◽  
...  
Keyword(s):  
Cell Types ◽  
Cytoplasmic Dynein ◽  
Opposite Polarity ◽  
Model System ◽  
Organelle Transport ◽  
Transport Activity ◽  
Microtubule Motors ◽  
Microtubule Motor ◽  
Dynactin Complex ◽  
Binding Subunit

Kinesin II is a heterotrimeric plus end–directed microtubule motor responsible for the anterograde movement of organelles in various cell types. Despite substantial literature concerning the types of organelles that kinesin II transports, the question of how this motor associates with cargo organelles remains unanswered. To address this question, we have used Xenopus laevis melanophores as a model system. Through analysis of kinesin II–mediated melanosome motility, we have determined that the dynactin complex, known as an anchor for cytoplasmic dynein, also links kinesin II to organelles. Biochemical data demonstrates that the putative cargo-binding subunit of Xenopus kinesin II, Xenopus kinesin II–associated protein (XKAP), binds directly to the p150Glued subunit of dynactin. This interaction occurs through aa 530–793 of XKAP and aa 600–811 of p150Glued. These results reveal that dynactin is required for transport activity of microtubule motors of opposite polarity, cytoplasmic dynein and kinesin II, and may provide a new mechanism to coordinate their activities.

scholarly journals Dynein Supports Motility of Endoplasmic Reticulum in the FungusUstilago maydis

2002 ◽  
Vol 13 (3) ◽  
pp. 965-977 ◽  
Author(s):  
Roland Wedlich-Söldner ◽  
Irene Schulz ◽  
Anne Straube ◽  
Gero Steinberg
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Intracellular Transport ◽  
Fluorescent Protein ◽  
Brefeldin A ◽  
Cytoplasmic Dynein ◽  
Organelle Transport ◽  
Minor Importance ◽  
Dependent Transport

The endoplasmic reticulum (ER) of most vertebrate cells is spread out by kinesin-dependent transport along microtubules, whereas studies in Saccharomyces cerevisiae indicated that motility of fungal ER is an actin-based process. However, microtubules are of minor importance for organelle transport in yeast, but they are crucial for intracellular transport within numerous other fungi. Herein, we set out to elucidate the role of the tubulin cytoskeleton in ER organization and dynamics in the fungal pathogen Ustilago maydis. An ER-resident green fluorescent protein (GFP)-fusion protein localized to a peripheral network and the nuclear envelope. Tubules and patches within the network exhibited rapid dynein-driven motion along microtubules, whereas conventional kinesin did not participate in ER motility. Cortical ER organization was independent of microtubules or F-actin, but reformation of the network after experimental disruption was mediated by microtubules and dynein. In addition, a polar gradient of motile ER-GFP stained dots was detected that accumulated around the apical Golgi apparatus. Both the gradient and the Golgi apparatus were sensitive to brefeldin A or benomyl treatment, suggesting that the gradient represents microtubule-dependent vesicle trafficking between ER and Golgi. Our results demonstrate a role of cytoplasmic dynein and microtubules in motility, but not peripheral localization of the ER inU. maydis.

scholarly journals Bidirectional organelle transport can occur in cell processes that contain single microtubules.

1985 ◽  
Vol 100 (1) ◽  
pp. 322-326 ◽  
Author(s):  
M P Koonce ◽  
M Schliwa
Keyword(s):  
Microscopic Study ◽  
High Voltage ◽  
Intracellular Transport ◽  
Model System ◽  
Force Generation ◽  
Organelle Transport ◽  
Electron Microscopic ◽  
Voltage Electron ◽  
Intracellular Organelle ◽  
Cell Processes

Intracellular organelle transport was studied in a new model system, the giant freshwater ameba Reticulomyxa. The ameba extends a large reticulate network of cytoplasmic strands in which various phase-dense organelles can be seen to move at a rate of up to 25 microns/s. This combined light and high voltage electron microscopic study shows that organelles move bidirectionally in even the finest network strands that contain only a single microtubule. In terms of microtubule-associated intracellular transport, this observation defines a minimum set of conditions necessary for such movement. The implications of this finding for possible models of force generation are discussed.

scholarly journals Opposite-polarity motors activate one another to trigger cargo transport in live cells

2009 ◽  
Vol 187 (7) ◽  
pp. 1071-1082 ◽  
Author(s):  
Shabeen Ally ◽  
Adam G. Larson ◽  
Kari Barlan ◽  
Sarah E. Rice ◽  
Vladimir I. Gelfand
Keyword(s):  
Intracellular Transport ◽  
Cytoplasmic Dynein ◽  
Opposite Polarity ◽  
Live Cells ◽  
Organelle Transport ◽  
S2 Cells ◽  
Drosophila Melanogaster S2 Cells ◽  
Cargo Transport ◽  
Bidirectional Transport ◽  
Necessary And Sufficient

Intracellular transport is typically bidirectional, consisting of a series of back and forth movements. Kinesin-1 and cytoplasmic dynein require each other for bidirectional transport of intracellular cargo along microtubules; i.e., inhibition or depletion of kinesin-1 abolishes dynein-driven cargo transport and vice versa. Using Drosophila melanogaster S2 cells, we demonstrate that replacement of endogenous kinesin-1 or dynein with an unrelated, peroxisome-targeted motor of the same directionality activates peroxisome transport in the opposite direction. However, motility-deficient versions of motors, which retain the ability to bind microtubules and hydrolyze adenosine triphosphate, do not activate peroxisome motility. Thus, any pair of opposite-polarity motors, provided they move along microtubules, can activate one another. These results demonstrate that mechanical interactions between opposite-polarity motors are necessary and sufficient for bidirectional organelle transport in live cells.

Microtubule Dynamics and Organelle Transport in Reticulomyxa

1990 ◽  
Vol 48 (3) ◽  
pp. 194-195
Author(s):  
Yih-Tai Chen ◽  
Ursula Euteneuer ◽  
Ken B. Johnson ◽  
Michael P. Koonce ◽  
Manfred Schliwa
Keyword(s):  
Plasma Membrane ◽  
Light Microscopy ◽  
Cytoplasmic Dynein ◽  
Living Cells ◽  
Microtubule Dynamics ◽  
Model Systems ◽  
Organelle Transport ◽  
Biochemical Characteristics ◽  
Dependent Transport

The application of video techniques to light microscopy and the development of motility assays in reactivated or reconstituted model systems rapidly advanced our understanding of the mechanism of organelle transport and microtubule dynamics in living cells. Two microtubule-based motors have been identified that are good candidates for motors that drive organelle transport: kinesin, a plus end-directed motor, and cytoplasmic dynein, which is minus end-directed. However, the evidence that they do in fact function as organelle motors is still indirect.We are studying microtubule-dependent transport and dynamics in the giant amoeba, Reticulomyxa. This cell extends filamentous strands backed by an extensive array of microtubules along which organelles move bidirectionally at up to 20 μm/sec (Fig. 1). Following removal of the plasma membrane with a mild detergent, organelle transport can be reactivated by the addition of ATP (1). The physiological, pharmacological and biochemical characteristics show the motor to be a cytoplasmic form of dynein (2).

Microtubule motors and other maps

1990 ◽  
Vol 48 (3) ◽  
pp. 1-1
Author(s):  
Richard B. Vallee
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Cytoplasmic Dynein ◽  
Organelle Transport ◽  
Structural Components ◽  
Microtubule Associated Proteins ◽  
Microtubule Motors ◽  
Associated Proteins ◽  
The Subject ◽  
Intracellular Motility

Microtubules are involved in a number of forms of intracellular motility, including mitosis and bidirectional organelle transport. Purified microtubules from brain and other sources contain tubulin and a diversity of microtubule associated proteins (MAPs). Some of the high molecular weight MAPs - MAP 1A, 1B, 2A, and 2B - are long, fibrous molecules that serve as structural components of the cytamatrix. Three MAPs have recently been identified that show microtubule activated ATPase activity and produce force in association with microtubules. These proteins - kinesin, cytoplasmic dynein, and dynamin - are referred to as cytoplasmic motors. The latter two will be the subject of this talk.Cytoplasmic dynein was first identified as one of the high molecular weight brain MAPs, MAP 1C. It was determined to be structurally equivalent to ciliary and flagellar dynein, and to produce force toward the minus ends of microtubules, opposite to kinesin.

scholarly journals Stable tug-of-war between kinesin-1 and cytoplasmic dynein upon different ATP and roadblock concentrations

2020 ◽  
Author(s):  
Gina A. Monzon ◽  
Lara Scharrel ◽  
Ashwin DSouza ◽  
Ludger Santen ◽  
Stefan Diez
Keyword(s):  
Cytoplasmic Dynein ◽  
Gliding Motility ◽  
Force Balance ◽  
Stochastic Simulations ◽  
Transport Systems ◽  
Organelle Transport ◽  
Atp Concentration ◽  
Dynein Motor ◽  
Bidirectional Movement

ABSTRACTThe maintenance of intracellular processes like organelle transport and cell division depend on bidirectional movement along microtubules. These processes typically require kinesin and dynein motor proteins which move with opposite directionality. Because both types of motors are often simultaneously bound to the cargo, regulatory mechanisms are required to ensure controlled directional transport. Recently, it has been shown that parameters like mechanical motor activation, ATP concentration and roadblocks on the microtubule surface differentially influence the activity of kinesin and dynein motors in distinct manners. However, how these parameters affect bidirectional transport systems has not been studied. Here, we investigate the regulatory influence of these three parameter using in vitro gliding motility assays and stochastic simulations. We find that the number of active kinesin and dynein motors determines the transport direction and velocity, but that variations in ATP concentration and roadblock density have no significant effect. Thus, factors influencing the force balance between opposite motors appear to be important, whereas the detailed stepping kinetics and bypassing capabilities of the motors have only little effect.

scholarly journals Peroxisomes move by hitchhiking on early endosomes using the novel linker protein PxdA

2015 ◽  
Author(s):  
John Salogiannis ◽  
Martin J. Egan ◽  
Samara L. Reck-Peterson
Keyword(s):  
Molecular Motors ◽  
Intracellular Transport ◽  
Coiled Coil ◽  
Leading Edge ◽  
Time Lapse ◽  
Early Endosome ◽  
Organelle Transport ◽  
The Novel ◽  
Coiled Coil Region ◽  
Early Endosomes

Eukaryotic cells use microtubule-based intracellular transport for the delivery of many subcellular cargos, including organelles. The canonical view of organelle transport is that organelles directly recruit molecular motors via cargo-specific adaptors. In contrast to this view, we show here that peroxisomes move by hitchhiking on early endosomes, an organelle that directly recruits the transport machinery. Using the filamentous fungus Aspergillus nidulans we find that hitchhiking is mediated by a novel endosome-associated linker protein, PxdA. PxdA is required for normal distribution and long-range movement of peroxisomes, but not early endosomes or nuclei. Using simultaneous time-lapse imaging we find that early endosome-associated PxdA localizes to the leading edge of moving peroxisomes. We identify a coiled-coil region within PxdA that is necessary and sufficient for early endosome localization and peroxisome distribution and motility. These results present a new mechanism of microtubule-based organelle transport where peroxisomes hitchhike on early endosomes and identify PxdA as the novel linker protein required for this coupling.

Rab3D directs organelle transport by recruiting cytoplasmic dynein motor protein Tctex-1 to secretory membranes of osteoclast

Bone ◽  
2009 ◽  
Vol 44 ◽  
pp. S160 ◽  
Author(s):  
N.J. Pavlos ◽  
J. Xu ◽  
H. Feng ◽  
P. Ng ◽  
T. Cheng ◽  
...  
Keyword(s):  
Motor Protein ◽  
Cytoplasmic Dynein ◽  
Organelle Transport ◽  
Dynein Motor

Regulation of the intracellular distribution of cytoplasmic dynein by serum factors and calcium

1993 ◽  
Vol 105 (2) ◽  
pp. 579-588 ◽  
Author(s):  
S.X. Lin ◽  
C.A. Collins
Keyword(s):  
Intracellular Transport ◽  
Cytoplasmic Dynein ◽  
Serum Starvation ◽  
Experimental Conditions ◽  
Extracellular Medium ◽  
Mitotic Apparatus ◽  
Cultured Fibroblasts ◽  
Serum Factors ◽  
Intracellular Organelles ◽  
Punctate Staining

Previous work has indicated that cytoplasmic dynein localizes primarily to lysosomes in cultured fibroblasts, consistent with a function for dynein in retrograde movement. We now show that dynein can be redistributed from a lysosome-associated pool to a more diffuse cytoplasmic pool upon shifting fibroblasts to culture medium lacking serum for several hours. This effect on dynein localization is readily reversed upon addition of serum, with a substantial return to a control appearance of punctate staining within 10 minutes. The serum effect appears to be selective for dynein, in that the localization of kinesin and the overall morphology of intracellular organelles does not change. However, the distribution of kinesin-positive vesicles and lysosomes does appear to be altered during serum starvation, in that these organelles are located to greater extents in the peripheral regions of the cell. Dynein is also associated with the mitotic apparatus, but this localization does not change in response to serum starvation. Removal of calcium from the extracellular medium also results in the loss of punctate dynein staining, which can be recovered upon addition of calcium to calcium-free medium. The redistribution of dynein observed under these experimental conditions may reflect the activity of a regulatory process controlling the association of dynein with organelles, thereby providing one means of modulating intracellular transport.

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