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At least 127 records · Page 7Linked to original sources
PubMed2006

A re-examination of the minor role of unstirred layers during the measurement of transport coefficients of Chara corallina internodes with the cell pressure probe.

The impact of unstirred layers (USLs) during cell pressure probe experiments with Chara corallina internodes has been quantified. The results show that the hydraulic conductivity (Lp) measured in hydrostatic relaxations was not significantly affected by USLs even in the presence of high water flow intensities ('sweep-away effect'). During pressure clamp, there was a reversible reduction in Lp by 20%, which was explained by the constriction of water to aquaporins (AQPs) in the C. corallina membrane and a rapid diffusional equilibration of solutes in arrays where water protruded across AQPs. In osmotic experiments, Lp, and permeability (Ps) and reflection (sigma s) coefficients increased as external flow rate of medium increased, indicating some effects of external USLs. However, the effect was levelling off at 'usual' flow rates of 0.20-0.30 m s(-1) and in the presence of vigorous stirring by air bubbles, suggesting a maximum thickness of external USLs of around 30 microm including the cell wall. Because the diameters of internodes were around 1 mm, internal USLs could have played a significant or even a dominating role, at least in the presence of the rapidly permeating solutes used [acetone, 2-propanol and dimethylformamide (DMF)]. A comparison of calculated (diffusion kinetics) and of measured permeabilities indicated an upper limit of the contribution of USLs for the rapidly moving solute acetone of 29%, and of 15% for the less rapidly permeating DME The results throw some doubt on recent claims that in C. corallina, USLs rather than the cell membrane dominate solute uptake, at least for the most rapidly moving solute acetone.

Chara↗
PubMed2004

Intracellular axial current in Chara corallina reflects the altered kinetics of ions in cytoplasm under the influence of light.

Recent experiments demonstrate that the concentration of Ca2+ in cytoplasm of Chara corallina internodal cells plays important role in electrical excitation of the plasma membrane. The concentration of free Ca2+ in the cytoplasm -[Ca2+]c is also sensitive to visible light. Both phenomena were simultaneously studied by noninvasive measuring action potential (AP) and magnetic field with a superconducting quantum interference device magnetometer in very close vicinity of electrically excited internodal C. corallina cells. A temporal shift in the depolarization maximum, which progressively occurred after transferring cells from the dark into the light, can be explained by the extended Othmer model. Assuming that the change in membrane voltage during the depolarization part of AP is the direct consequence of an activation of [Ca2+]c sensitive Cl- channels, the model simulations compare well with the experimental data. We can say that we have an example of electrically elicited AP that is of biochemical nature. Electric and magnetic measurements are in good agreement.

Action Potentials↗
PubMed2005

Spatial coordination of chloroplast and plasma membrane activities in Chara cells and its disruption through inactivation of 14-3-3 proteins.

In Chara corallina cells exposed to continuous light, external pH (pH(o)) and photosystem II (PSII) photochemical yield show correlated banding patterns. Photosynthetic activity is low in cell regions producing alkaline zones and high in the acid regions. We addressed the question whether (and how) photosynthetic activity and plasma membrane (PM) H+-pumping and H+-conductance are coupled in the different bands. First, PM H+-pump activity was stimulated with fusicoccin. This resulted in a more acidic pH in the acid bands without disturbing the correlation of photosynthetic electron transport and H+ fluxes across the PM. Next, H+-pump activity was reduced through microinjection of a phosphorylated peptide matching the canonical 14-3-3 binding motif RSTpSTP in the acid cell region. Microinjection induced a rapid (~5 min) rise in pH(o) by ca. 1.0 unit near the injection site, whereas the injection of the non-phosphorylated peptide had no effect. This pH rise confirms the supposed inhibition of the H+-pump upon the detachment of 14-3-3 proteins from the H+-ATPase. However, the PSII yield in the cell regions corresponding to the new alkaline peak remained high, which violated the normal inverse relations between the pH(o) and PSII photochemical yield. We conclude that the injection of the competitive inhibitor of the H+-ATPase disrupts the balanced operation of PM H+-transport and photosynthetic electron flow and promotes electron flow through alternative pathways.

14-3-3 Proteins↗
PubMed2005

GA3 content in young and mature antheridia of Chara tomentosa estimated by capillary electrophoresis.

The content of gibberellic acid (GA3) in male sex organs of Chara tomentosa L. was estimated using capillary electrophoresis. Young antheridia contained 0.25 microg GA3 while mature ones 0.48 microg per antheridium. Although there are significant differences in GA3 content in antheridia between C. vulgaris and C. tomentosa, these values calculated per one spermatid are 2.4 and 3.3 pg, respectively. The present results compiled with the previous knowledge about regulation of GA3-dependent development of Characeae species allow an implication that the mechanisms controlling antheridia differentiation in both species can be similar.

Animals↗
PubMedDate not supplied

[Modeling of hysteresis in pH pattern formation along the cell membrane of algae Chara corallina].

It is known that illumination of the algae Chara corallina results in the formation along the membrane of regions with inhomogeneous distribution of pH. It was shown that, in a particular range of illumination intensities, two states with different pH distribution are realized at one and the same value of light intensity: an entirely homogeneous state and completely formed structures (pattern). The transition from the homogeneous state to the pattern formation takes place at one value of light intensity, and the back transition, at another light intensity, i.e., the hysteresis is observed. This phenomenon was studied by mathematical modeling. The mechanism of hysteresis is discussed.

Cell Membrane↗
PubMed1982

Investigation of the effects of continuous-wave, pulse- and amplitude-modulated microwaves on single excitable cells of Chara corallina.

Single internodal excitable cells of Chara corallina were exposed to CW, pulse-modulated and sinusoidally modulated S-band microwave fields in a temperature-controlled waveguide exposure chamber. All electrical measurements were made external to the waveguide (ie, under no impressed microwave field). The dependent variables measured before, during, and after exposure to the S-band microwave fields included: resting potential, amplitude of the action potential, rise and decay time of the action potential, conduction velocity, and excitability. Cells maintained at 22 +/- 0.1 degrees C during exposure showed no consistent or statistically significant microwave-dependent alterations in any of the dependent variables.

Chlorophyta↗
PubMed1996

Partial characterization of a putative 110 kDa myosin from the green alga Chara corallina by in vitro binding of fluorescent F-actin.

Using the binding of heterologous, rhodamine phalloidin-labelled F-actin in vitro, two F-actin binding proteins were identified in protein extracts from the green alga Chara corallina after fractionation by anion exchange chromatography. The first protein, a putative myosin, released laterally bound F-actin at ATP-concentrations as low as 1 microM; equivalent concentrations of ADP were not effective. Binding of F-actin was inhibited by the sulfhydryl-alkylating agent N-ethylmaleimide (NEM). Binding of F-actin was also abolished by a monoclonal anti-myosin (J14) previously used for immunodetection and immunolocalization in internodal cells (Grolig et al., 1988, Eur J Cell Biol 47: 22-31). Immunoblotting with J14 detected a 110 kDa polypeptide only in those protein fractions that had revealed ATP-sensitive binding of F-actin. The putative myosin bound with mediocre affinity to immobilized calmodulin and free Ca(2+)-concentration made no difference to this binding affinity. In contrast to the putative myosin, the second, less abundant protein revealed ATP-insensitive and end-wise binding to the microfilament and was not recognized by the anti-myosin antibody.

Actins↗
PubMed1998

ANTHERIDIAL CHROMATIN CONDENSATION FACTOR FROM MALE SEX ORGANS OF CHARA TOMENTOSA.

Incubation of roots (Melandrium noctiflorum) in crude extracts obtained from maturing male sex organs of Chara tomentosa results in supercondensation of mitotic chromosomes. The observed changes, including a number of 'secondary effects', confirm precisely with all those kinds of alterations which are typical of M-phase cells in antheridial filaments during their progression from the proliferative period of development towards terminal differentiation into sperm cells. Biological assays performed with proteinaceous extracts of maturing antheridia indicate that the activity is confined merely to a fraction containing low molecular (4.5 kDa) peptides, termed as ACCF (antheridial chromatin condensation factor). Microchemical tests showed the acidic character of ACCF and its apparent ability to form cross-links with histones and DNA-histone complexes, mediated presumably by disulfide bridges. The characteristic punctate distribution of vesicles within the cytoplasm suggests the internalization of rhodamine-isothiocyanate labelled ACCF (ACCF-TRITC) by endocytosis, implying possible sequestering and nuclear translocation upon release from the endosomal/lysosomal compartment of the cell. Copyright 1998 Academic Press.

Journal Article↗
PubMed1995

Myosin from alga Chara: unique structure revealed by electron microscopy.

Cytoplasmic streaming is a fundamental process for the transport of molecules and organelles in plant cells. A myosin-like protein responsible for cytoplasmic streaming in fresh water alga, Chara corallina, has been purified. Electron micrographs of this protein revealed that it has two heads with similar size and shape to those of muscle myosin (myosin II) but its tail was shorter than that of muscle myosin with a globular structure at its distal end like that of myosin V. The myosin cosedimented with vesicles composed of phosphatidyl serine like other non-muscle myosin species involved in intracellular transport.

Actins↗
PubMed2001

Functional expression of a chimeric myosin-containing motor domain of Chara myosin and neck and tail domains of Dictyostelium myosin II.

We succeeded in expressing a chimeric myosin that comprises the motor domain of characean algal myosin, (the fastest known motor protein), and the neck and tail domains of Dictyostelium myosin II. Although the chimeric myosin showed an ATPase activity comparable to that of muscle myosin (15 times higher than that of the wild-type Dictyostelium myosin II), the motile activity of the chimera was only 1.3 times higher than that of the wild-type. However, this is the first chimeric myosin that showed motile activity faster than at least one of the parent myosins. It was suggested, therefore, that the motor domain of Chara myosin has the potential for performing fast sliding movement.

Actins↗
PubMed1995

Micromanipulation of statoliths in gravity-sensing Chara rhizoids by optical tweezers.

Infrared laser traps (optical tweezers) were used to micromanipulate statoliths in gravity-sensing rhizoids of the green alga Chara vulgaris Vail. We were able to hold and move statoliths with high accuracy and to observe directly the effects of statolith position on cell growth in horizontally positioned rhizoids. The first step in gravitropism, namely the physical action of gravity on statoliths, can be simulated by optical tweezers. The direct laser microirradiation of the rhizoid apex did not cause any visible damage to the cells. Through lateral positioning of statoliths a differential growth of the opposite flank of the cell wall could be induced, corresponding to bending growth in gravitropism. The acropetal displacement of the statolith complex into the extreme apex of the rhizoid caused a temporary decrease in cell growth rate. The rhizoids regained normal growth after remigration of the statoliths to their initial position 10-30 micrometers basal to the rhizoid apex. During basipetal displacement of statoliths, cell growth continued and the statoliths remigrated towards the rhizoid tip after release from the optical trap. The resistance to statolith displacement increased towards the nucleus. The basipetal displacement of the whole complex of statoliths for a long distance (>100 micrometers) caused an increase in cell diameter and a subsequent regaining of normal growth after the statoliths reappeared in the rhizoid apex. We conclude that the statolith displacement interferes with the mechanism of tip growth, i.e. with the transport of Golgi vesicles, either directly by mechanically blocking their flow and/or, indirectly, by disturbing the actomyosin system. In the presence of the actin inhibitor cytochalasin B the optical forces required for acropetal and basipetal displacement of statoliths were significantly reduced to a similar level. The lateral displacement of statoliths was not changed by cytochalasin B. The results indicate: (i) the viscous resistance to optical displacement of statoliths depend mainly on actin, (ii) the lateral displacement of statoliths is not impeded by actin filaments, (iii) the axially directed actin-mediated forces against optical displacement of statoliths (for a distance of 10 micrometers) are stronger in the basipetal than in the acropetal direction, (iv) the forces acting on single statoliths by axially oriented actin filaments are estimated to be in the range of 11-110 pN for acropetal and of 18-180 pN for basipetal statolith displacements.

Actins↗
PubMed1994

Surface potentials near the mouth of the large-conductance K+ channel from Chara australis: a new method of testing for diffusion-limited ion flow.

The kinetics of single K+ channels were derived for patch-clamp recordings of membrane patches excised from cytoplasmic drops from the plant, Chara australis R. Br. Specifically, the "tilt effect" model of MacKinnon, Latorre and Miller (1989. Biochemistry 28:8092-8099) has been used to measure the electrostatic potential (surface PD) and fixed charge at the entrances of the channel. The surface PD is derived from the difference between the trans-pore potential difference (PD) and that between the two bulk phases. The trans-pore PD is probed using three voltage-dependent properties of the channel. These are (1) the association and dissociation rates of Ca2+ binding to the channel, from both the cytoplasmic and vacuolar solutions. These were determined from the mean blocked and unblocked durations of the channel in the presence of either 20 mmol liter-1 vacuolar or 1 mmol liter-1 cytoplasmic Ca2+; (2) the closing rate of the channel's intrinsic gating process. This was determined from the mean channel open time in the absence of vacuolar Ca2+ at membrane PDs more negative than -100 mV; and (3) the effect of Mg2+ on channel conductance when added to solutions initially containing 3 mmol liter-1 KCl. The voltage dependence of properties 1 and 2 shifts along the voltage axis according to the ionic strength of the bathing media, consistent with the presence of negative charge in the channel vestibules. Furthermore, the magnitude of this shift depends on the current in a manner consistent with diffusion-limited ion flow in the channel (i.e., the rate of ion diffusion in the external electrolyte limits the channel conductance). Mg2+ on either side of the membrane alters channel conductance in a voltage-dependent way. A novel feature of the Mg2+ effect is that it reverses, from a block to an enhancement, when the membrane PD is more negative than -70 mV. This reversal only appears in solutions of low ionic strength. The attenuating effect is due to voltage-dependent binding of Mg2+ within the pore, which presumably plugs the channel. The enhancing effect is due to screening by Mg2+ of surface potentials arising from diffusion-limited flow of K+. All experimental approaches give a consistent picture of K+ permeation in which the surface charge and convergence permeability of the cytoplasmic vestibule are the major factors in determining channel conductance. The cytoplasmic vestibule has a charge density of -0.035 C/m2 which is similar to that found for maxi K channels in rat muscle.(ABSTRACT TRUNCATED AT 400 WORDS)

Biological Transport↗
PubMed1994

Cl- and K+ channel currents during the action potential in Chara. Simultaneous recording of membrane voltage and patch currents.

Patch currents in the cell attached-mode and action potentials (AP) have been recorded simultaneously in internodal cells of Chara corallina. The action potentials are closely correlated with transient patch currents. With pipettes containing either 50 mM CaCl2 or 100 mM KCl plus 1 or 5 mM CaCl2, these transients measured up to 100 to 200 pA per patch at zero mV. Transients had a mean duration (time during which the current was > or = half maximum peak amplitude) of about 1 sec, a maximum slope for current rising of about 400 pA sec-1 and a maximum rate of about 100 pA sec-1 for current decay, with no obvious effect of external Ca2+ on either of these parameters. In well-resolved recordings of current transients triggered by an action potential (AP), activities of two types of Cl--conducting channels (15 and 38 pS) have been identified. Since activity of these channels was only observed during action potentials but not upon positive voltage steps, these channels are not directly voltage gated but point to a cytoplasmic gating factor which accumulates during excitation and propagates from excited areas to the patch. A K(+)-conducting channel (40 pS) could be identified as well during an AP, when 100 mM KCl was in the pipette solution. The activity of this channel relaxed at the end of the APs with a time constant of about 3 sec. Stimulated activity of this channel is understood to cause the repolarization overshoot during the final phase of the action potential, whereas the transient activation of the Cl- channels determines the fast voltage changes of the action potential.

Action Potentials↗
PubMed1993

Enzyme kinetics of the prime K+ channel in the tonoplast of Chara: selectivity and inhibition.

The prime potassium channel from the tonoplast of Chara corallina has been analyzed in terms of an enzymatic kinetic model (Gradmann, Klieber & Hansen 1987, Biophys. J. 53:287) with respect to its selectivity for K+ over Rb+ and to its blockage by Cs+ and by Ca2+. The channel was investigated by patch-clamp techniques over a range of membrane voltages (Vm, referred to an extracytoplasmic electrical potential of zero) from -200 mV to +200 mV under various ionic conditions (0 to 300 mM K+, Rb+, Cs+, Ca2+, and Cl-) on the two sides of isolated patches. The experimental data are apparent steady-state current-voltage relationships under all experimental conditions used and amplitude histograms of the seemingly noisy open-channel currents in the presence of Cs+. The used model for K+ uniport comprises a reaction cycle of one binding site through four states, i.e., (1) K(+)-loaded and charged, facing the cytoplasm, (2) K(+)-loaded and charged facing the vacuole, (3) empty, facing the vacuole, and (4) empty, facing the cytoplasm. Vm enters the system in the form of a symmetric Eyring barrier between state 1 and 2. The numerical results for the individual rate constants are (in 10(6)s-1 for zero voltage and 1 M substrate concentration): k12: 1,410, k21: 3,370, k23: 105,000, k32: 10,600, k34: 194, k43: 270, k41: 5,290, k14: 15,800. For the additional presence of an alternate transportee (here Rb+), the model can be extended in an analog way by another two states ((5) Rb(+)-loaded and charged, facing cytoplasm, and (6) Rb(+)-loaded and charged, facing vacuole) and six more rate constants (k45: 300, k54: 240, k56: 498, k65: 4,510, k63: 4,070, k36: 403). This six-state model with its unique set of fourteen parameters satisfies the complete set of experimental data. If the competing substrate can be bound but not translocated (here Cs+ and Ca2+). k56 and k65 of the model are zero, and the stability constants Kcyt (= k36/k63) and Kvac (= k45/k54) turn out to be Kcyt(Ca2+): 250 M-1 x exp(Vm/(64 mV)), kvac(Ca2+): 10 M-1 x exp(-Vm/(66 mV)), Kcyt(Cs+): 0, and Kvac(Cs+): 46 M-2 x exp(-Vm/(12.25 mV)).(ABSTRACT TRUNCATED AT 400 WORDS)

Calcium↗
PubMed1992

Chara plasmalemma at high pH: voltage dependence of the conductance at rest and during excitation.

The high pH state of Chara plasmalemma (Bisson, M.A., Walker, N.A. 1980. J. Membrane Biol. 56:1-7) was investigated to obtain detailed current-voltage (I/V) and conductance-voltage (G/V) characteristics in the pH range 7.5 to 12. The resting conductance started to increase at a pH as low as 8.5, doubling at pH 9.5, but the most notable increases occurred between pH 10.5 and 11.5, as observed previously (Bisson, M.A., Walker, N.A. 1980. J. Membrane Biol. 56:1-7; Bisson, M.A., Walker, N.A. 1981. J. Exp. Bot. 32:951-971). The slopes (and shapes) of the I/V curves varied even over minutes, suggesting a shifting population of open channels. Possible contributions of the permeabilities to H+ and OH-, PH and POH, respectively, to the increase in membrane conductance were calculated in the pH range 8.5 to 12. If PH is the main cause for the increase in conductance, it would have to rise by three orders of magnitude between pH 8.5 and 11.5, implying an enormous increase in the open-channel population as pH rises. On the other hand, a comparatively constant POH over that pH range would result in an increase in conductance due to the rise of OH- concentration. This indicates unchanging open-channel population. The transient excitation conductances at pH 7.5 and 11.5 were compared at a range of membrane PD (potential difference) levels. At more positive PD levels (near 0) the transient conductances showed little change as pH was increased. However, near the excitation threshold the conductance at high pH was slower to reach peak and its amplitude was diminished compared to that at neutral pH. This effect was found to be partially due to the pH change itself and partially due to less negative membrane PD at high pH. The changes in excitation transients developed gradually as pH of the medium was increased. These findings are discussed with a recent model of excitation in mind (Shiina, T., Tazawa, M. 1988. J. Membrane Biol. 106:135-139).

Action Potentials↗
PubMed1988

DNA-polymerase activity detected in situ. Relationship between incorporation of 3H-deoxytriphosphates and 3H-actinomycin D binding during the cell cycle in antheridial filaments of Chara vulgaris L. Effect of auxin and cytokinins.

DNA-polymerase activity during the cell cycle (S + G2 + M + C type) in antheridial filaments cells of Chara vulgaris was studied using the autoradiographic method. Incorporation of 3H-deoxytriphosphates (3H-dTPs) during the whole of interphase indicates, that the cell cycle is not accompanied by distinct changes in enzyme activity. Incorporation of 3H-dTPs was also observed in spermatids and in early stages of spermatogenesis. Intensity of 3H-dTPs incorporation during interphase and spermatogenesis is similar to the intensity of 3H-actinomycin D (3H-AMD) binding. Auxin (IAA) and kinetin stimulate both 3H-AMD binding and 3H-dTPs incorporation; benzyladenine does not affect any of these processes. The in situ autoradiographic method of detecting DNA-polymerase activity reveals availability of DNA template for the enzyme rather than DNA polymerase activity itself.

Autoradiography↗
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↗
PubMed1993

Centrifugation causes adaptation of microfilaments: studies on the transport of statoliths in gravity sensing Chara rhizoids.

The actin cytoskeleton is involved in the positioning of statoliths in tip growing Chara rhizoids. The balance between the acropetally acting gravity force and the basipetally acting net outcome of cytoskeletal force results in the dynamically stable position of the statoliths 10-30 micrometers above the cell tip. A change of the direction and/or the amount of one of these forces in a vertically growing rhizoid results in a dislocation of statoliths. Centrifugation was used as a tool to study the characteristics of the interaction between statoliths and microfilaments (MFs). Acropetal and basipetal accelerations up to 6.5 g were applied with the newly constructed slow-rotating-centrifuge-microscope (NIZEMI). Higher accelerations were applied by means of a conventional centrifuge, namely acropetally 10-200 g and basipetally 10-70 g. During acropetal accelerations (1.4-6g), statoliths were displaced to a new stable position nearer to the cell vertex (12-6.5 micrometers distance to the apical cell wall, respectively), but they did not sediment on the apical cell wall. The original position of the statoliths was reestablished within 30 s after centrifugation. Sedimentation of statoliths and reduction of the growth rates of the rhizoids were observed during acropetal accelerations higher than 50 g. When not only the amount but also the direction of the acceleration were changed in comparison to the natural condition, i.e., during basipetal accelerations (1.0-6.5 g), statoliths were displaced into the subapical zone (up to 90 micrometers distance to the apical cell wall); after 15-20 min the retransport of statoliths to the apex against the direction of acceleration started. Finally, the natural position in the tip was reestablished against the direction of continuous centrifugation. Retransport was observed during accelerations up to 70 g. Under the 1 g condition that followed the retransported statoliths showed an up to 5-fold increase in sedimentation time onto the lateral cell wall when placed horizontally. During basipetal centrifugations > or = 70 g all statoliths entered the basal vacuolar part of the rhizoid where they were cotransported in the streaming cytoplasm. It is concluded that the MF system is able to adapt to higher mass accelerations and that the MF system of the polarly growing rhizoid is polarly organized.

Acceleration↗