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PubMed2005

Removal of the nitro and phenyl groups from NPPB decreases its inhibitory effect on cytoplasmic streaming in the alga Nitella hookeri.

Structural analogues of the arylaminobenzoate 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB), prepared using a simple reductive amination sequence, were tested for their effects on cytoplasmic streaming rates in the alga Nitella hookeri. Cytoplasmic streaming was sensitive to NPPB, with an IC50 value of 24 micromol/L. Removal of the nitro group from the benzoate ring decreased the IC50 to 455 micromol/L. The introduction of an extra carbon or double bond into the aliphatic chain had no effect on activity. Loss of the phenyl group decreased potency, with an IC50 of 6.4 mmol/L. These data are the first documenting the relative inhibitory effects of structural changes to arylaminobenzoates in algae. Patch-clamp data and the effects of tetrapentyl ammonium chloride on streaming suggest that the nitro and phenyl groups may act by inhibiting both K+ and Cl- channels. This is likely, through changes in the membrane potential, to affect Ca2+ fluxes and action potentials, thereby slowing cytoplasmic streaming.

Aminobenzoates↗
PubMed2005

Dynein and the actin cytoskeleton control kinesin-driven cytoplasmic streaming in Drosophila oocytes.

Mass movements of cytoplasm, known as cytoplasmic streaming, occur in some large eukaryotic cells. In Drosophila oocytes there are two forms of microtubule-based streaming. Slow, poorly ordered streaming occurs during stages 8-10A, while pattern formation determinants such as oskar mRNA are being localized and anchored at specific sites on the cortex. Then fast well-ordered streaming begins during stage 10B, just before nurse cell cytoplasm is dumped into the oocyte. We report that the plus-end-directed microtubule motor kinesin-1 is required for all streaming and is constitutively capable of driving fast streaming. Khc mutations that reduce the velocity of kinesin-1 transport in vitro blocked streaming yet still supported posterior localization of oskar mRNA, suggesting that streaming is not essential for the oskar localization mechanism. Inhibitory antibodies indicated that the minus-end-directed motor dynein is required to prevent premature fast streaming, suggesting that slow streaming is the product of a novel dynein-kinesin competition. As F-actin and some associated proteins are also required to prevent premature fast streaming, our observations support a model in which the actin cytoskeleton triggers the shift from slow to fast streaming by inhibiting dynein. This allows a cooperative self-amplifying loop of plus-end-directed organelle motion and parallel microtubule orientation that drives vigorous streaming currents and thorough mixing of oocyte and nurse-cell cytoplasm.

Actins↗
PubMedDate not supplied

Entamoeba motility: dynamics of cytoplasmic streaming, locomotion and translocation of surface-bound particles, and organization of the actin cytoskeleton in Entamoeba invadens.

The dynamics of cytoplasmic streaming, retrograde translocation of externally bound particles and locomotion by Entamoeba invadens were compared. Locomoting amoebae were monopodial, exhibited fountain flow cytoplasmic streaming and translocated externally bound erythrocytes to the rear of cells. The rates of rearward flow of peripheral cytoplasmic vacuoles and of the externally bound particles were equal to the rate of cell forward locomotion. Rhodamine-phalloidin staining revealed a distinct cortical polymerized actin cytoskelton. This was least evident about the periphery of the advancing pseudopod, increased in density toward the rear of the cell and was most concentrated in the uroid. A monoclonal anti-eucaryotic actin antibody, which recognized monomeric Entamoeba actin on immunoblots, stained trophozoites by indirect immunofluorescence throughout the cytoplasm, but not in the cortical regions stained by rhodamine-phalloidin. This and other evidence implied that the antibody recognized only unpolymerized actin in Entamoeba. We propose that locomotion, cytoplasmic streaming and translocation of externally bound particles are driven by a common actin-based mechanism in Entamoeba, possibly involving retrograde cortical actin flow and recycling.

Actins↗
PubMed1994

Variation in velocity of cytoplasmic streaming and gravity effect in characean internodal cells measured by laser-Doppler-velocimetry.

Velocities of cytoplasmic streaming were measured in internodal cells of Nitella flexilis L. and Chara corallina Klein ex Willd. by laser-Doppler-velocimetry to investigate the possibility of non-statolith-based perception of gravity. This was recently proposed, based on a report of gravity-dependent polarity of cytoplasmic streaming. Our measurements revealed large spatial and temporal variation in streaming velocity within a cell, independent of the position of the cell with respect to the direction of gravity. In 58% of the horizontally positioned cells the velocities of acropetal and basipetal streaming, measured at opposite locations in the cell, differed significantly. In 45% of these, basipetal streaming was faster than acropetal streaming. In 60% of the vertically positioned cells however the difference was significant, downward streaming was faster in only 61% of these. When cell positions were changed from vertical to horizontal and vice versa the cells reacted variably. A significant difference between velocities in one direction, before and after the change, was observed in approx. 70% of the measurements, but the velocity was faster in the downward direction, as the second position, in only 70% of the significantly different. The ratio of basipetal to acropetal streaming velocities at opposite locations of a cell was quite variable within groups of cells with a particular orientation (horizontal, normal vertical, inverted vertical). On average, however, the ratio was close to 1.00 in the horizontal position and approx. 1.03 in the normal vertical position (basipetal streaming directed downwards), which indicates a small direct effect of gravity on streaming velocity. Individual cells, however, showed an increased, as well as a decreased, ratio when moved from the horizontal to the vertical position. No discernible effect of media (either Ca(2+)-buffered medium or 1.2% agar in distilled water) on the streaming velocities was observed. The above mentioned phenomenon of graviperception is not supported by our data.

Actins↗
PubMed1990

Movement of vesicles in cytoplasmic streaming in plasmodium.

The moving velocities of vesicles in the cytoplasmic streaming of a slime mold were measured, in which all of the vesicles passing through a designated window were counted. Vesicles in the streaming are distributed in their moving velocities and the distribution itself varies with time. The mean velocity of vesicles and its standard deviation were found to exhibit a linear relationship, suggesting a possibility that vesicles in the cytoplasm would also be involved in force generation.

Cytoplasmic Streaming↗
PubMed1991

Cytoplasmic streaming in Chara rhizoids: studies in a reduced gravitational field during parabolic flights of rockets.

In-vivo videomicroscopy of Chara rhizoids under 10(-4)g demonstrated that gravity affected the velocities of cytoplasmic streaming. Both, the acropetal and basipetal streaming velocities increased on the change to microgravity. The endogenous difference in the velocities of the oppositely directed cytoplasmic streams was maintained under microgravity, yet the difference was diminished as the basipetal streaming velocity increased more than the acropetal streaming velocity. Direction and structure of microfilaments labeled by rhodamine-phalloidin had not changed after 6 min of microgravity.

Acceleration↗
PubMed2000

The role of plant villin in the organization of the actin cytoskeleton, cytoplasmic streaming and the architecture of the transvacuolar strand in root hair cells of Hydrocharis.

In many types of plant cell, bundles of actin filaments (AFs) are generally involved in cytoplasmic streaming and the organization of transvacuolar strands. Actin cross-linking proteins are believed to arrange AFs into the bundles. In root hair cells of Hydrocharis dubia (Blume) Baker, a 135-kDa polypeptide cross-reacted with an antiserum against a 135-kDa actin-bundling protein (135-ABP), a villin homologue, isolated from lily pollen tubes. Immunofluorescence microscopy revealed that the 135-kDa polypeptide co-localized with AF bundles in the transvacuolar strand and in the sub-cortical region of the cells. Microinjection of antiserum against 135-ABP into living root hair cells induced the disappearance of the transvacuolar strand. Concomitantly, thick AF bundles in the transvacuolar strand dispersed into thin bundles. In the root hair cells, AFs showed uniform polarity in the bundles, which is consistent with the in-vitro activity of 135-ABP. These results suggest that villin is a factor responsible for bundling AFs in root hair cells as well as in pollen tubes, and that it plays a key role in determining the direction of cytoplasmic streaming in these cells.

Actins↗
PubMed1976

The migrating thermodynamic quantum hypothesis for cytoplasmic streaming, sodium pumping and other cell biological phenomena, deduced from biofunctional considerations of the ultrastructure of brush border microvilli.

An attempt is made to reconcile experimental data dealing with, inter alia, cytoplasmic streaming in Characean algae, contraction in actomyosin systems. Na+- and -K+-simtulated ATPase activity and the ultrastructure of brush border microvilli. It is postulated that myosin molecules transfer energy from ATP to an actin-containing filament and that a high energy conformation is subsequently propagated along the filament. At regularly spaced intervals corresponding to the length of an actin-tropomyosin subunit, the propagation of high energy involves rejection of a pressure pulse in the direction of cytoplasmic streaming. Proteins in solution capable of storing the thermodynamic energy represented by the pressure pulse will either migrate in the opposite direction or conserve the quantized cytoplasmic flow generated by the actin-containing filaments. At sites where actin filaments are attached to the plasma membrane the high energy is propagated in another direction leading to expulsion of sodium ions and neutralization of the vectorial pressure pulse.

Actomyosin↗
PubMed1988

The mechanism of cytoplasmic streaming in characean algal cells: sliding of endoplasmic reticulum along actin filaments.

Electron microscopy of directly frozen giant cells of characean algae shows a continuous, tridimensional network of anastomosing tubes and cisternae of rough endoplasmic reticulum which pervade the streaming region of their cytoplasm. Portions of this endoplasmic reticulum contact the parallel bundles of actin filaments at the interface with the stationary cortical cytoplasm. Mitochondria, glycosomes, and other small cytoplasmic organelles enmeshed in the endoplasmic reticulum network display Brownian motion while streaming. The binding and sliding of endoplasmic reticulum membranes along actin cables can also be directly visualized after the cytoplasm of these cells is dissociated in a buffer containing ATP. The shear forces produced at the interface with the dissociated actin cables move large aggregates of endoplasmic reticulum and other organelles. The combination of fast-freezing electron microscopy and video microscopy of living cells and dissociated cytoplasm demonstrates that the cytoplasmic streaming depends on endoplasmic reticulum membranes sliding along the stationary actin cables. Thus, the continuous network of endoplasmic reticulum provides a means of exerting motive forces on cytoplasm deep inside the cell distant from the cortical actin cables where the motive force is generated.

Actins↗
PubMed2002

Visualization of peroxisomes in living plant cells reveals acto-myosin-dependent cytoplasmic streaming and peroxisome budding.

Here we examine peroxisomes in living plant cells using transgenic Arabidopsis thaliana plants expressing the green fluorescent protein (GFP) fused to the peroxisome targeting signal 1 (PTS1). Using time-lapse laser scanning confocal microscopy we find that plant peroxisomes exhibit fast directional movement with peak velocities approaching 10 microm s(-1). Unlike mammalian peroxisomes which move on microtubules, plant peroxisome movement is dependent on actin microfilaments and myosin motors, since it is blocked by treatment with latrunculin B and butanedione monoxime, respectively. In contrast, microtubule-disrupting drugs have no effect on peroxisome streaming. Peroxisomes were further shown to associate with the actin cytoskeleton by the simultaneous visualization of actin filaments and peroxisomes in living cells using GFP-talin and GFP-PTS1 fusion proteins, respectively. In addition, peroxisome budding was observed, suggesting a possible mechanism of plant peroxisome proliferation. The strong signal associated with the GFP-PTS1 marker also allowed us to survey cytoplasmic streaming in different cell types. Peroxisome movement is most intense in elongated cells and those involved in long distance transport, suggesting that higher plants use cytoplasmic streaming to help transport vesicles and organelles over long distances.

Actin Cytoskeleton↗
PubMed2004

Contribution of the actomyosin motor to the temperature-dependent translational diffusion of water by cytoplasmic streaming in Elodea canadensis cells.

The extent to which the actomyosin motor responsible for cytoplasmic streaming contributes to the translational diffusion of water in Elodea canadensis cells was studied by a nuclear magnetic resonance (NMR) spin-echo technique. The relative contribution of the actomyosin motor was determined from the corresponding apparent diffusion coefficient by the Einstein-Smolukhovsky relation. It is equal to the difference between the diffusional displacements of the cytoplasmic and the bulk water (deltaX). The NMR data show that the temperature dependence of deltaX is humpshaped, which is characteristic of enzyme reactions. At the same time, the apparent diffusion coefficient of cytoplasmic water increases with an increase in temperature. The most significant contribution of the actomyosin motor to deltaX is observed at temperatures below 20 degrees C. Within the temperature range of 20 to 33 degrees C, deltaX changes only slightly, and a further increase in temperature reduces deltaX to zero.

Actomyosin↗
PubMedDate not supplied

[Study of cytoplasm streaming as a cytophysiological method in radiolabeled experiment].

Estimation of influence of ionizing radiation, high-frequency electromagnetic radiation and their combined action on a higher water plant Elodea canadensis has been carried out using cytophysiological method of determination of the cytoplasm streaming rate. It was shown that low-intensive electromagnetic radiation modifies reaction of the differentiated cells on radiolesion. The rate of cytoplasm streaming can be used as an informative characteristic of plant cell state in radiobiological experiment.

Cobalt Radioisotopes↗
PubMed2003

Ca2+-dependent cessation of cytoplasmic streaming induced by hypertonic treatment in Vallisneria mesophyll cells: possible role of cell wall-plasma membrane adhesion.

In mesophyll cells of the aquatic angiosperm Vallisneria gigantea Graebner, a rapid and transient inhibition of cytoplasmic streaming was induced by hypertonic treatment with sorbitol. Higher concentrations of sorbitol induced the response more rapidly and in more cells. The response to hypertonic treatment was strictly dependent on the presence of extracellular Ca2+ and was sensitive to Ca2+-channel blockers, including the stretch-activated Ca2+-channel blocker Gd3+. Deplasmolyzed cells never responded to a second hypertonic treatment administered immediately after plasmolysis and subsequent deplasmolysis. Responsiveness was gradually recovered during 24 h of incubation; however, cycloheximide, cordycepin, and trypsin completely suppressed the recovery. Although an Arg-Gly-Asp (RGD) hexapeptide markedly disturbed the pattern of cytoplasmic streaming, it exhibited no specific effects on the response to hypertonic treatment or on the recovery of responsiveness. Taken together, our results demonstrate that leaf mesophyll cells in a multicellular plant can respond to mechanical stimuli and that a Ca2+ influx through stretch-activated Ca2+ channels plays an indispensable role in the response. Furthermore, the possible involvement of RGD-insensitive but trypsin-sensitive protein factor(s), whose function is impaired by detachment of the plasma membrane from the cell wall, is suggested.

Calcium↗
PubMed1990

Cooperative regulation of cytoplasmic streaming and ca fluxes by pfr and photosynthesis in vallisneria mesophyll cells.

In mesophyll cells of Vallisneria gigantea Graebner, Ca(2+) regulates the induction and cessation of cytoplasmic streaming. Streaming is induced when the level of calcium in the cytoplasm is lowered through light-accelerated release of Ca(2+) from the cells (S Takagi, R Nagai [1988] Plant Physiol 88: 228-232). We have now initiated an investigation on the nature of the photoreceptor(s) that are involved in the regulation of Ca(2+) movements across the cell membrane and of streaming. Streaming is induced only when phytochrome exists in the phytochrome-far redabsorbing form (Pfr)-and photosynthesis is allowed to take place for at least 4 minutes. The former effect is typically photoreversible by red and far-red light, and phytochrome is spectro-photometrically detectable in the crude extract from the leaves. The latter effect is assessed in terms of the wavelength dependency and the effects of diuron and atrazine, two inhibitors of photosynthesis. A similar requirement for Pfr and photosynthesis is found to be associated with the acceleration of Ca(2+) efflux in the protoplasts. The results suggest that phytochrome and photosynthetic pigment(s) cooperatively regulate cytoplasmic streaming via modulation of the Ca(2+) transport in the cell membrane.

Journal Article↗
PubMed2000

The pattern of acropetal and basipetal cytoplasmic streaming velocities in Chara rhizoids and protonemata, and gravity effect on the pattern as measured by laser-Doppler-velocimetry.

The spatial pattern of acropetal and basipetal cytoplasmic streaming velocities has been studied by laser-Doppler-velocimetry (LDV) in the positively gravitropic (downward growing) rhizoids of Chara globularis Thuill. and for the first time in the negatively gravitropic (upward growing) protonemata. The LDV method proved to be precise and yielded reproducible results even when tiny differences in velocities were measured. In the apical parts of the streaming regions of both cell types, acropetal streaming was faster than basipetal streaming. Starting at the apical reversal point of streaming, the velocity increased basipetally with the distance from that point and became fairly constant close to the basal reversal point; subsequently, the velocity decreased slightly acropetally as the apical reversal point was again approached. There was no change in velocity at the basal reversal point. However, at the apical reversal point there was an abrupt decrease in velocity. The pattern of the ratio of acropetal to basipetal streaming velocity (VR) was a function of the relative distance of the site of measurement from the apical reversal point rather than a function of the absolute distance. Upon inversion of the rhizoids, the VR decreased on average by 3.8% (+/- 0.4%), indicating that the effect of gravity on the streaming velocity was merely physical and without a physiological amplification. Rhizoids that had developed on the slowly rotating horizontal axis of a clinostat, and had never experienced a constant gravity vector, were similar to normally grown rhizoids with respect to VR pattern. In protonemata, the VR pattern was not significantly different from that in rhizoids although the direction of growth was inverse. In rhizoids, oryzalin caused the polar organization of the cell to disappear and nullified the differences in streaming velocities, and cytochalasin D decreased the velocity of basipetal streaming slightly more than that of acropetal streaming. Cyclopiazonic acid, known as an inhibitor of the Ca2+-ATPase of the endoplasmic reticulum, also reduced the streaming velocities in rhizoids, but had slightly more effect on the acropetal stream. It is possible that the endogenous difference in streaming velocities in both rhizoids and protonemata is caused by differences in the cytoskeletal organization of the opposing streams and/or loading of inhibitors (like Ca2+) from the apical/subapical zone into the basipetally streaming endoplasm.

Cytochalasin D↗
PubMed1976

Photon correlation analysis of cytoplasmic streaming.

Laser light scattering has been used to investigate particle movements in a plant cell. Intensity autocorrelation functions are obtained by digital photon correlation of laser light scattered from cells of Nitella opaca both during cytoplasmic streaming and during the transitory cessation of streaming induced by electrical stimulation. The average velocity computed from the periodic oscillation in the intensity autocorrelation function during streaming corresponds to the velocity estimated using light microscopy. An estimate of the distribution of streaming velocities has been obtained from the decay in the amplitude of the envelope of the autocorrelation function derived from a streaming cell.

Cytoplasm↗
PubMed2004

Cytoplasmic streaming in plants.

Plant cells are surrounded by a cell wall composed of polysaccharides and hence can change neither their form nor their position. However, active movement of organelles (cytoplasmic streaming or protoplasmic streaming) is observed in plant cells, and involvement of the actin/myosin system in these processes has been suggested. Successful biochemical and biophysical approaches to studying myosins have extensively promoted the understanding of the molecular mechanism underlying these phenomena.

Actin Cytoskeleton↗
PubMed1986

The role of microfilaments in cytoplasmic streaming in Drosophila follicles.

During the last phase of oogenesis in Drosophila, nurse cell cytoplasm can be seen to be streaming into the growing oocyte when visualized in time-lapse films. This process can be reversibly inhibited by cytochalasins. The distribution of F-actin filaments in the nurse cells has been studied by staining with rhodamine-conjugated phalloidin. At the beginning of cytoplasmic streaming (stage 10B) increasingly thick bundles of microfilaments formed, many of which spanned the nurse cell cytoplasm from the cell membrane to the nuclear membrane. The association of F-actin with the nuclear membrane persisted when nurse cell nuclei were isolated mechanically. The experimental evidence suggests that microfilament contraction in the nurse cells leads to cytoplasmic streaming by pressure flow.

Actin Cytoskeleton↗