Calcium gradients in tip growing plant cells visualized by chlorotetracycline fluorescence

With chlorotetracycline (CTC)-fluorescence a tip-to-base Ca²⁺ gradient is visualized in all tested, tip-growing plant cells: pollen tubes of Lilium longiflorum, root hairs of Lepidium sativum, moss caulonema of Funaria hygrometrica, fungal hyphae of Achlya and in the alga Acetabularia mediterranea. The fluorescence gradients in the different species vary in intensity and extension. Sometimes a punctate mobile CTC-fluorescence, in the size range of mitochondria, is observed. Bursting cells lose their fluorescence rapidly, indicating a cytoplasmic localization of the gradient. Only in Acetabularia is the wall also fluorescent with CTC. The results are interpreted as evidence for a general role of a calcium gradient in tip growth.Abbreviation CTC chlorotetracycline     

PIP kinases and their role in plant tip growing cells

Phosphatidylinositol (4,5) bisphosphate, [PtdIns(4,5)P2], is a signaling lipid involved in many important processes in animal cells such as cytoskeleton organization, intracellular vesicular trafficking, secretion, cell motility, regulation of ion channels, and nuclear signaling pathways. In the last years PtdIns(4,5)P2 and its synthesizing enzyme, phosphatidylinositol phosphate kinase (PIPK), has been intensively studied in plant cells, revealing a key role in the control of polar tip growth. Analysis of the PIPK members from Arabidopsis thaliana, Oryza sativa and Physcomitrella patens showed that they share some regulatory features with animal PIPKs but also exert plant-specific modes of regulation. This review aims at giving an overview on the PIPK family from Arabidopsis thaliana and Physcomitrella patens. Even though their basic structure, modes of activation and physiological role is evolutionary conserved, modules responsible for plasma membrane localization are distinct for different PIPKs, depending on differences in physiological and/or developmental status of cells, such as polarized and non-polarized.     

Calcium influx at the tip of growing root-hair cells of Arabidopsis thaliana

The role of extracellular Ca²⁺ in root-hair tip growth has been investigated in Arabidopsis thaliana (L.) Heynh. Root-hair length was found to be dependent on the concentration of Ca²⁺ in the growth medium, with maximum length achieved at [Ca²⁺] of 0.3–3.0 mM. Using a non-intrusive calcium-specific vibrating microelectrode, an extracellular Ca²⁺ gradient was detected at the tips of individual growing root-hair cells. The direction of the gradient indicated a net influx of Ca²⁺ into root-hair cells. No gradient was detected near the sides of the root hairs or at the tips of non-growing root hairs. When root hairs were exposed to the Ca²⁺-channel blocker nifedipine, tip growth stopped and the extracellular Ca²⁺ gradient was abolished. These results indicate that Ca²⁺ influx through plasma-membrane Ca²⁺ channels is required for normal root-hair tip growth.Abbreviation APW artificial pond water We thank L.F. Jaffe, W. Kuhtreiber and A. Miller of the National Vibrating Probe Facility, Marine Biological Laboratory, Woods Hole, Mass., USA for their technical assistance and helpful discussions. We also thank Liam Dolan, Martin Steer, and Susan Ford for helpful discussions. This research was supported by National Science Foundation grant PCM-9004568.     

Microtubules spatial alterations in root cells of Brassica rapa under clinorotation

Organization of tubulin cytoskeleton in epidermis and cortex cells in different root growth zones in Brassica rapa L. 6-day-old seedlings under clinorotation has been investigated. It was shown that changes in cortical microtubules orientation occur only in the distal elongation zone. In control, cortical microtubule arrays oriented transversely to the root long axis. Whereas under clinorotation an appearance of shorter randomly organized cortical microtubules was observed. Simultaneously, a significant decrease in a cell length in the central elongation zone under clinorotation was revealed. It is suggested that the decline of anisotropic growth, typical for central elongation zone cells, is connected with cortical microtubules disorientation under clinorotation.     

Structural changes of chloroplasts and galactolipid contents in Chlorella cells during clinorotation

Research of chloroplast ultrastructure in Chlorella cells grown during long-term period under clinorotation has been carried out. Different changes of the chloroplast structure, concerning both the amount of starch grains and stroma electron density as well as membrane system have been revealed. Occurrence of more significant bends of the thylakoids compared to the control and more loose arrangement of the thylakoids in a bunch were noted. The most significant changes were observed in a membrane system, in particular, appearance of the non-uniform expansions of the inter- and intrathylakoid spaces in Chlorella chloroplasts. Taking into account the role of galactolipids as important components of the photosynthetic membranes, the content of general galactolipids, monogalactosyldiacylglycerol and digalactosyldiacylglicerol in Chlorella cells was determined. It was assumed that statistically significant increase of the galactolipid content, especially of MDG, can probably be one of the reasons of membrane system reorganizations in Chlorella cells under altered gravity.     

Cell cycle changes in transformed cells growing under serum-free conditions.

Polyoma transformed hamster cells (PyBHK) and SV40 transformed mouse cells (SV3T3) were transferred in culture using crystalline trypsin followed by neutralisation with soybean trypsin inhibitor. Such cells were able to proliferate freely in defined medium without any serum supplement and without any intervening period of adaptation. However, growth rates were reduced under serum-free conditions. Re-establishment of rapid growth rates could be achieved by addition of serum, with the rate attained being proportional to the serum concentration. Irrespective of the prevailing rates of growth, percentages of cells synthesising DNA were the same. However, the rate at which DNA was being synthesised was found to change proportionately with the changes in overall growth rate.     

Calcium and rhythms in plant cells

Plants cells, like any other living organism, experience the daily rotation of the Earth. They also depend strikingly on light, as a result of which much of the plant's biochemistry, physiology, and behaviour are temporally organised with respect to the environmental oscillation of day and night. Here we review the most recent findings on plants rhythms and how they seem to be so tightly connected to calcium-signalling aspects. We also try to establish parallels between different cell types, such as pollen tubes and fungal hyphae, where the existence and function of rhythms and oscillations is not obvious. Additionally, we discuss new methodologies and how these are shaping our current working hypothesis to study Ca²⁺ rhythms in plant cells.     

Cytoskeleton orientation in epidermis cells of roots generated de novo in leaf explants under clinorotation

The anatomy, cytoskeleton orientation, and thickness of the cell wall of the root growth zones generated de novo in vitro under clinorotation (simulated microgravity) were studied. The anatomical structure of the roots generated de novo from the cambium cells of the leaf explant petiole is similar to the structure of embryonic roots. The root cell differentiation in vitro during the clinorotation does not differ from the control in main features. Changes in the tubulin cytoskeleton orientation under clinorotation were detected in the epidermis of distal elongation zone (that is apparently associated with specific physiological properties of the cells in this zone). A tendency towards the thinning of the root cell walls in vitro under conditions of simulated microgravity was established.     

Calcium channels and signal transduction in plant cells

An increasing number of studies indicate that changes in cytosolic free Ca²⁺ ([Ca²⁺]_c) mediate specific types of signal transduction in plant cells. Modulation of [Ca²⁺]_c is likely to be achieved through changes in the activity of Ca²⁺ channels, which catalyse passive influx of Ca²⁺ to the cytosol from extracellular and intracellular compartments. Voltage-sensitive Ca²⁺ channels have been detected in the plasma membranes of algae, where they control membrane electrical properties and cell turgor. These channels are sensitive to 1,4-dihydropyridines, which in animal cells specifically affect one class of voltage-regulated plasma membrane Ca²⁺ channel. Ca²⁺-permeable channels with different pharmacological properties have been found in the plasma membrane of higher plants. Recent evidence suggests the existence of two discrete classes of Ca²⁺ channel co-resident in the vacuolar membrane (tonoplast) of higher plants. The first is gated by inositol 1,4,5-trisphosphate, and bears a number of similarities to its animal counterpart which is located in the endoplasmic reticulum (ER). The second tonoplast Ca²⁺ channel is voltage-operated. However, the specific roles of these tonoplast channels in signal transduction have yet to be elucidated.     

Ultrastructure of statocytes and cells of distal elongation zone of Arabidopsis thaliana under clinorotation

Results of the electron-microscopic investigation of root apices of Arabidopsis thaliana 3-, 5- and 7-days-old seedlings grown in the stationary conditions and under clinorotation are presented. It was shown the similarity in the root apex cell ultrastructure in control and under clinorotation. At the same time there were some differences in the ultrastructure of statocytes and the distal elongation zone under clinorotation. For the first time the sensitivity of ER-bodies, which are derivatives of GER and contain beta-glucosidase, to the influence of simulated microgravity was demonstrated by increased quantity and area of ER-bodies at the cell section as well as by higher variability of their form under clinorotation. A degree of these changes correlated with the duration of clinorotation. On the basis of experimental data a protective role of ER-bodies in adaptation of plants to microgravity is supposed.     

Transport changes in synchronously growing CHO and L cells

A reduced rate of transport of 2-amino-isobutyric acid (AIB), was found to be associated with M and early G₁ periods in synchronized populations of CHO and L cells. A doubling of the transport rate occurred with progress of CHO cells further into G₁, and the specific transport activity remained constant during the rest of the cycle, The rate of uridine and thymidine transport also doubled at this point in the cell cycle. There was no change in Km values. The rise in transport rate was blocked by cycloheximide, or when the cells were prevented from leaving metaphase by colcemid.     

Proton pumps of the vacuolar membrane in growing plant cells

Plant growth results from the division, enlargement and specialization of cells. The two processes of the enlargement and the differentiation of cells are not spatially separated in plant tissue. We focus our attention here on the enlargement and elongation of cells. In most cases, growing plant cells contain a large central vacuole. The acid growth theory is based on the space-filling function of the large vacuole. The active transport systems in the vacuolar membrane are essential for maintenance of high osmotic pressure and for the expansion of the vacuole. The secondary active transport systems of the vacuole for sugars and ions are driven by the proton-motive force which is generated by the vacuolar H⁺-ATPase and H⁺-translocating inorganic pyrophosphatase. In this review, the relationship between cell elongation and these enzymes of the vacuolar membrane is emphasized.     

The case for multinet growth in growing walls of plant cells

The basis of multinet gwoth, the multinet growth hypothesis, is examined in view of recent criticisms. It is shown that the strain across a growing wall may be calculated by simple means and the expected reorientations are deduced (a) for a wall in which the microfibrils of the innermost wall lamella always lie helically with the same pitch and (b) in which the microfibrils lie at random. Calculations are presented both for cells increasing in length only and for cells also increasing in breadth. Both the strains and the reorientations are smaller than commonly implied and are too small to be reliably detectable in wall sections. Observations on wall sections cannot therefore be accepted as proof that microfibril reorientation has not occured and it is concluded that the multinet growth hypothesis still stands as applying both to parenchyma and to collenchyma cells. In view of the wide dispersity in the structure of the walls of growing cells, it is recommended that the qualifying multinet should be dropped and replaced by passive reorientation.Abbreviation MGH multinet growth hypothesis     

Adenilate pool and thylakoid ATP-synthase content in pea leaves under clinorotation

It is known that plant resistance to stress factors is connected with energy metabolism. The energy stored in the process of photophosphorylation in the form of ATP is used then to support respiration, transpiration, organic compound synthesis, growth and development as well as to restore cell structure after its damage under extremal environmental factors. Transformation of light energy into chemical energy of ATP is catalyzed by thylakoid membrane enzymatic complex of ATPsynthase-CF1CF0. Its activity and amount in the thylakoid membrane depends on plant growth conditions. The aim of this work was investigation of clinorotation effect on light-induced dynamics of adenyl nucleotides (AMP, ADP and ATP) and estimation of CF1CF0 content in thylakoids of pea leaves grown under slow clinorotation and vertical control.     

Postural control under clinorotation in upside-down catfish, Synodontis nigriventris.

The upside-down catfish Synodontis nigriventris has a unique habit of swimming and resting upside-down in free water. This behavior leads to the assumption that the catfish has a specific gravity information processing system. We examined the postural control behaviors in the catfish under clinorotation which is usually used for producing pseudo-microgravity. Synodontis nigriventris kept its body posture at a stable area of the rotated flask in which the catfish was kept, when it was clinorotated at the rate of 60 rpm. In contrast to Synodontis nigriventris, a related species, Corydoras paleatus, did not show such steady postural control. When the flask was rotated at a lower rate of 30 rpm or a higher rate of 100 rpm, Synodontis nigriventris as well as Corydoras paleatus showed a considerable disturbed control of body posture. In this condition, they were frequently rotated with the flask. These findings suggest that Synodontis nigriventris has a high ability to keep upside-down posture and the gravity sensation in this catfish is likely to contribute to its different postural control from that of many other fishes.     

Effect of monensin on mineral balance in growing ruminants reared under different environmental temperatures

To test the effect of monensin on the mineral balance of growing cattle under different environmental temperatures, 24 male steers were assigned in a 2 × 2 factorial arrangement, contrasting 0 and 85 mg monensin/animal per day at 24.3 and 33.2 °C (environmental temperatures). Monensin effect was directly modulated by the environmental temperature: it increased apparent retentions of P (P=0.066), Na (P=0.005) and K (P=0.003), at the higher temperature and decreased these apparent retentions at the lower temperature, as compared with non-supplemented animals. Monensin increased fecal Ca (P=0.037), and urinary P (P=0.002), Na (P=0.003), K (P=0.014), Mg (P=0.051) and Zn (P=0.091), with higher concentrations of these minerals in animals held at 24.3 °C and lower concentrations in those at 33.2 °C, as compared with non-supplemented animals. Monensin decreased serum Mg (P=0.001) and increased serum Zn (P=0.071) in animals at 33.2 °C and increased serum Mg and decreased serum Zn at 24.3 °C. Irrespective of temperature, monensin increased both apparent absorption (P=0.058) and apparent retention (P=0.093) of P, and also urine Cu (P=0.085). Environmental temperature modulated monensin effects on mineral balance. Monensin increased apparent retention of several minerals in animals under heat stress.