NaCl-induced amoeboid plastids and mitochondria in meristematic cells of barley roots

The barley (Hordeum vulgare L.) seeds were germinated in the non-saline conditions after 12 h imbibition in 2 % of NaCl solution. The results of treatment were: (l) the membrane system in meristematic cells of root tips developed well; (2) many profiles of endoplasmic reticulum and Golgi bodies appeared; and (3) the quantities of amoeboid plastids and amoeboid mitochondria increased. Thus the inhibitory effects of short-term NaCl stress on plants were reversible, and simultaneously NaCl treatment enhanced the metabolic activities in cells. The amoeboid form may be an adaptive form of plastids or mitochondria to an enhanced metabolic activity.     

Distribution of ubiquitin protein in meristematic mesophyll ceils of barley leaves

The distribution of ubiquitin protein in meristematic mesophyll cells of barley (Hordeum vulgare L.) leaves was investigated by using immunofluorescence microscopy. Simultaneous observation of nuclei was achieved byDAPI (4 6-diamidino-2-phenylindol-dihydrochloride) staining. A strong correlation between the chromatin organisation and the ubiquitin distribution could be observed. Interphase nuclei revealed an intense content of ubiquitin and accumulation of ubiquitin at the nuclear envelope, whereas condensed chromosomes of dividing cells excluded any ubiquitin appearance. During cell division, the aggregation of ubiquitin protein was detected in the area of the mitotic spindle in anaphase as well as the area of the cell plate in the late telophase.     

Temperature-induced absorbance changes in developing barley chloroplasts

Absorbance changes on cooling and heating of barley ( Hordeum vulgare L. cv. IB65) chloroplasts greened for 12, 48 and 72 h were investigated to understand the structural changes during biogenesis of chloroplast membranes. Upon cooling the chloroplast suspension from 24 to 8°C, a positive absorbance change occurred at 678, 435 and 495 nm in 12, 48 and 72 h greened chloroplasts. During heating from 24 to 45°C negative absorbance changes were observed with some shifts in positions in different chloroplast preparations and a simultaneous increase in absorbance between 690 and 735 nm. For chloroplasts developed for 12, 48 and 72 h the changes in absorbance on cooling were 3.8, 3.3 and 1.9% at 678 nm, and on heating, 8.9, 8.3 and 4.1% at 680 nm.
The differences in absorbance changes are considered as an indication of variations in the structural organization and composition of developing chloroplasts. The reversibility of the absorbance changes was maximum in chloroplasts greened for 72 h and minimum in chloroplasts greened for 12 h. This would suggest that fully developed chloroplasts have more flexibility towards temperature-induced changes in the membranes.     

Translocation of N to and from barley roots: its dependence on local nitrate supply in split-root culture

The relationship between availability of external nitrate and N translocation between root and shoot was studied in N-limited barley ( Hordeum vulgare L. cv. Golf). Nitrate-N was added at a relative rate (i.e. N added per unit time and unit N in plant biomass) of (1.09 da-¹, and distributed between the subroots at ratios of 50:50 or 80:20. The plants were grown for 13 days under these conditions of nitrate nutrition, and for another three days with the nitrate distribution reversed from 80:20 to 20:80. The nitrale-N doses thus experienced by individual subroots ranged from 2 to 11 mg N g-¹ root dry weight day-¹ . ¹⁵N-Nitrate labellings were performed after 2 to 3 and 12 to 13 days of nitrate nutrition. and 2 to 3 days after reversal of nitrate additions. For all treatments, between 60 and 82% of the absorbed label initially left the root, and between 25 and 55% of the label recovered in roots had been supplied (cycled) via the shoot. Labelling of xylem N at the end of the 24-h labelling period ranged from to 36 to 46% indicating that a substantial fraction of the N in the xylem had been absorbed by the plant prior to labelling. It is concluded that cycling of N to roots, and cycling of N in the plant as a whole, is substantial also during N-limited growth. N allocation to roots increased with external nitrate dose. An increased utilization of non-translocated N as well as an increased translocation of N from the shoot contributed to this effect. Thus, the results indicate that increased external availability of N also increased the sink strength of the root for cycling N.     

Salinity induced nuclear and DNA degradation in meristematic cells of soybean (Glycine max (L.)) roots

Changes in the nuclei of meristematic root cells of soybean (Glycine max (L.) Merr. cv. Acme) in response to severe salinity were studied. Root growth was inhibited by 200 mM NaCl, when 1 mM CaCl_2 was present in the culture media. Increasing CaCl_2 up to 5 mM partially prevented this inhibition. However, inhibition also occurred with 100~mM NaCl without CaCl_2. We examined the meristematic cells under a series of NaCl treatments. Nuclear deformation of the cells occurred with 24 h of 150 mM or higher NaCl, and was followed by degradation of nuclei in the apical region of the root. TEM observation and agarose gel electrophoretic analysis confirmed that root tip nuclear DNA deformed or degraded with 150 mM or higher NaCl concentrations.     

Soluble polypeptides from meristematic and mature cells of Allium cepa L. roots

By using electrophoresis on SDS polyacrylamide gels we have studied the soluble protein fraction from a section of the meristem, where most cells are in the division last cycle, and from the mature region of Allium cepa L. roots. In order to estimate the apparent rate of synthesis of these polypeptides we labeled a series of roots with [¹⁴C]leucine and another with [³H]leucine. Coelectrophoresis was carried out by using polypeptides from both regions, their mol.wt. being between 20,000 and 100,000 daltons. The results show that most of the polypeptides in the soluble fraction are constantly present in a cell throughout its development. These constant polypeptides are synthesized at a high rate in the meristematic region. In the mature cells these polypeptides show only a low labeling rate, while a small number of specific polypeptides appear to have a very high rate of metabolism.     

Localization of phytosiderophore release and of iron uptake along intact barley roots

Under iron deficiency the release of so-called phytosiderophores by roots of barley plants ( Hordeum vulgare L. cv. Europa) was greater by a factor of 10 to 50 compared to iron-sufficient plants. This enhanced release occurred particularly in apical zones of the seminal roots and in the lateral root zones. Under iron deficiency, uptake rates for iron, supplied as FeIII phytosiderophore, increased by a factor of ca 5 as compared to iron-sufficient plants. This enhanced uptake rate for iron was also much more pronounced in apical than in basal root zones. In contrast, with supply of the synthetic iron chelate, FelII EDDHA (ferric diaminoethane-N, N-di- o -hydroxyphenyl acetic acid), the Fe deficiency-enhanced uptake rates for iron were only small and similar along the roots, except for the lateral root zones. The high selectivity of barley roots for uptake and translocation of FeIII phytosiderophores compared with FeIII EDDHA is reflected by the fact that, at the same external concentration (2 μ M ), rates of uptake and translocation of iron from FeIII phytosiderophores were between 100 (Fe-sufficient) and 1 000 times higher (Fe-deficient plants) than from FeIII EDDHA. The relatively high rates of uptake and particularly of translocation of iron supplied as FeIII EDDHA in the zone of lateral root formation strongly suggest an apoplastic pathway of radial transport of the synthetic iron chelate into the stele in this root zone.
The results demonstrate that apical root zones are the main sites both for Fe deficiency-enhanced release of phytosiderophores and for uptake and translocation of iron supplied as FeIII phytosiderophores.     

Time-dependent changes in polypeptide and translatable mRNA levels caused by NaCl in barley roots

The effect of salt stress on polypeptide and translatable mRNA levels was examined in roots of barley ( Hordeum vulgare L. cv. CM 72). A salt-shock time course (200 m M NaCl added to the nutrient solution of 5-day old seedlings for up to 24 h) and a 6-day salt treatment (seedlings grown in nutrient solution containing 200 m M NaCl for 6 days) were compared. Roots of intact seedlings were labeled in vivo with [³⁵S]-methionine. Poly(A)⁺ RNA was isolated and assayed in an in vitro translation system. The changes in the levels of the labeled polypeptides and translation products were analyzed following separation on two-dimensional polyacrylamide gels. Over the salt-shock time course, the majority of the changes in polypeptides and translation products were quantitative, as were the changes at 6 days. Qualitative changes occurred during the salt-shock time course, but were not observed at 6 days. The levels of polypeptides and translation products differed at each point of the salt-shock time course and at 6 days. In addition, changes were observed at the shortest time points examined, indicating a rapid response to salt.     

Ultrastructural changes in aging leaves of a light-grown achlorophyllous mutant of barley

The pattern and sequence of cellular degradation during the course of leaf senescence remains obscure and the nature of the trigger that induces cell senescence is unknown. In order to probe the pre-mortem phase of senescence temporal changes in cell ultrastucture were studied in aging leaves of light-grown achlorophyllous Hordeum vulgare L. cv. Dyan mutant seedlings. Electron microscope examination of the ultrastructure of mesophyll cell plastids revealed the absence of ribosomes and a highly disorganized prolamellar body. Both the number and size of plastoglobuli increased with aging and this change coincided with depletion of starch grains and dilation of lamellar membranes. Aging of mesophyll cells occurred coincident with a decline in ribosome content of the cytoplasm and loss of matrix granularity. Loss of ribosomes associated with the outer nuclear envelope membrane and a reduction in chromatin were also apparent. Only after 10 days was there evidence of loss of internal membrane integrity and swelling of mitochondrial cristae. Compartmentation was thus maintained during the aging process with membrane dissolution occurring late in senescence. These results suggest that an inability to produce chlorophyll and carotenoids and form thylakoid stacks due to the absence of plastid ribosomes, contributes to the rapid onset of senescence in light-grown achlorophyllous seedlings. Furthermore, disruption of chloroplast ribosome synthesis/assembly may constitute part of the plastid signal involved in triggering cell senescence.     

Transport-related amino acid metabolism in germinating barley grains

When eight [¹⁴C]-labelled amino acids were separately injected into the endosperm of germinating (4 days at 20°C) barley (Hordeum vulgare L. cv. Himalaya) grains, the label was rapidly taken up by the scutellum and further transported to the shoot and roots. Some of the amino acids (leucine, lysine and asparagine) were transported in an intact form through the scutellum to the seedling, whilst glutamic acid and aspartic acid were largely converted to glutamine in the scutellum. Proline was mainly transported unchanged, but a small part of the label appeared in glutamine. Arginine was mostly broken down in the scutellum, possibly providing ammonia for the synthesis of glutamine. During further transport in the seedling there was a partial transfer of label from glutamine to asparagine, particularly in the shoot. None of the amino acids used supplied carbon for the synthesis of sucrose, glucose or fructose. Glutamine synthetase activity was particularly high in the scutellum during the period of rapid amino acid transport.     

Contrasting phosphate acquisition of mycorrhizal fungi with that of root hairs using the root hairless barley mutant

Comparisons between plant species or cultivars differing in root hair length have indicated a major impact of root hairs on the mycorrhizal dependency of plants with respect to phosphate (P) uptake. The current study aimed to investigate this relationship by comparing directly the mycorrhizal dependency of a spontaneous root hairless mutant, brb , in Hordeum vulgare cv Pallas and its wild type. Both brb and wild type were grown at different soil P levels in association with different mycorrhizal fungi. P uptake of brb and wild type was similar at high P levels, but P uptake by non-mycorrhizal brb plants at low P levels was substantially lower than that of the non-mycorrhizal wild-type plants. However, P uptake of the mutant was much increased by mycorrhizas and with one fungus, the additional P uptake was effectively translated into increased plant growth. Roots of the mutant contained typical colonization structures and a radioactive tracer confirmed P transport by the extraradical mycelium. This is the first direct evaluation of the relative effectiveness of root hairs and mycorrhizas. Mycorrhizas effectively substituted root hairs in P uptake, whereas the additional P was most often used less effectively in promoting plant growth than P provided by root hairs.     

Zinc-induced vacuolation in root meristematic cells of Festuca rubra L.

Abstract. The effect of Zn on vacuole development in root meristematic cells was examined in three cultivars of Festuca rubra: a Zn-tolerant cultivar (Merlin), a salt-tolerant cultivar (Hawk) and a non-tolerant cultivar (S.59) in order to determine whether or not vacuole development was related to Zn tolerance. Treatment with Zn greatly increased the percentage of cells in the apical meristem which were vacuolated in all three cultivars (7.01-fold increase in Merlin, 3.61-fold increase in Hawk and 5.39-fold increase in S.59 over the range 0–0.5 μg Zn cm^?3). Morphometric analyses on electromicrographs indicated that the percentage total vacuolar volume fraction of meristematic tissue was also increased by Zn treatment. Most of this increase was due to an increase in a particular component of the vacuole which was 0.2–0.5 μm in diameter, spherical or ovoid in outline and possessed a distinct amorphous electron dense matrix (Type A intravacuolar body). X-ray microanalysis revealed that this matrix was rich in calcium and phosphorus in control plants. In Zn-treated roots, elevated levels of Zn were found in the matrix in Merlin and Hawk, but not in S.59. In addition, intravacuolar membranous whorls or myelin bodies were more highly organized in Zn-treated Merlin and Hawk but not S.59. Elevated Zn levels were not found in any other vacuolar component nor in nuclei, nucleoli, cell walls or ground cytoplasm. Control roots of Merlin possessed a greater type A vacuolar volume fraction than either Hawk or S.59 which may confer a greater capacity to compartmentalize Zn at the onset of exposure to toxic metals than the other cultivars. Predictably, the EM procedures resulted in loss of Zn from the roots to the fixative and dehydrating solutions. However, the rate of loss was the same (approximately 75–80%) regardless of cultivar. These findings are discussed in relation to their possible role in the mechanisms of Zn tolerance in these cultivars.     

Apparent transinhibition of peptide uptake in the scutellum of barley grain

The uptake of glycylsarcosine (Gly-Sar) into scutella separated from germinating grains of barley ( Hordeum vulgare L. cv. Himalaya) is inhibited by other peptides; in most cases the inhibition is not purely competitive but of a mixed type (simultaneous increase in the apparent K_m and decrease in V_max) (Sopanen, T. 1979. FEBS Lett. 108: 447–450). The aim of the present experiments was to elucidate the mechanism of the mixed inhibition by studying how peptides already taken up into the cells affect the uptake of Gly-Sar.
When scutella were preincubated in the presence of various peptides, 11 of the 13 peptides tested inhibited the subsequent uptake of Gly-Sar by 10 to 45%. The inhibition, studied in detail with leucylleucine and prolylproline, was due to a decrease in V_max. The two peptides having no effect were glycylglycine and D-alanyl-L-alanine which are the only peptides known to date acting as purely competitive inhibitors when present together with the substrate Gly-Sar.
Preincubation with leucine, proline and alanine was not inhibitory, although preincubation with the corresponding dipeptides was. This result, together with the demonstration of intact leucylleucine in the scutella after preincubation with leucylleucine, indicates that the inhibition was caused by the intact peptides.
The results support the notion that in the mixed type inhibition the increase in the apparent K_m is due to competition for the carrier at the outside of the membrane, while the decrease in V_max is due to peptides taken up and binding to the carrier at the inside of the membrane.     

Effect of moderate salinity on patterns of potassium, sodium and chloride accumulation in cells near the root tip of barley: Role of differentiating metaxylem vessels

X-Ray microanalysis of fully hydrated, bulk-frozen samples was used to measure concentrations of potassium, sodium and chloride in various cell types along seminal roots of barley ( Hordeum vulgare L. cv. California Mariout) seedlings (1 to 150 mm from the tip). In the cytoplasm of all meristematic cells 1 mm from the root tip, the average concentrations of potassium and chloride were ca 200 and 15 m M , respectively. The potassium level was also high in the vacuoles of incipient xylem elements and did not drop to significantly lower values until 10 mm from the tip in protoxylem, 50 mm in early metaxylem and 150 mm in late metaxylem (LMX). Light microscopy observations (Nomarski optics) of hand-cut sections showed the presence of cytoplasmic strands and also the presence of intact cross walls in LMX up to a distance of 100 mm. Both quantitative analysis of ion contents and structural observations suggested that LMX elements act as a large transitional sink of accumulated ions and therefore may not function as a main pathway of transport until perforation of the end wall takes place 100–150 mm from the root tip. Treatment with 50 m M NaCl resulted in higher concentrations of sodium and chloride in LMX elements than in the surrounding cells, suggesting that living xylem elements, which develop a large central vacuole at an early stage of root differentiation, may assist in alleviating salinity stress in the meristematic region of barley root tips. Further, it is proposed that reabsorption of sodium and chloride from the LMX, especially before the disappearance of the cross walls, may provide a means of salinity tolerance.     

Effects of combined NaCl and CaCl2 salinity on photosynthetic parameters of barley grown in nutrient solution

The changes caused by NaCl− and CaCl₂-induced salinity on several leaf parameters have been measured in two cultivars of barley ( Hordeum vulgare L.) growing in a growth chamber in nutrient solution. Salinity was induced by adding to the nutrient solution equal weights of NaCl and CaCl₂, to obtain conductivities of 2, 6, 12, 19 and 26 dS m⁻¹. Salinity induced decreases in the leaf water potential and in the osmotic potential. Salinity did not induce significant changes in the relative photosynthetic pigment composition of barley leaves, the photosynthetic pigment stoichiometry for neoxanthin:violaxanthin cycle pigments:lutein:β-carotene:Chl b :Chl a being close to 3:6:14:12:25:100 (mol:mol). Salinity per se did not induce interconversions in the carotenoids within the violaxanthin cycle in most barley leaves. The PSII photochemistry of most barley leaves was unchanged by salinity. However, some apparently healthy leaves growing in high salinity exhibited sudden decreases in PSII photochemistry and increases in zeaxanthin (at the expense of violaxanthin), that preceded rapid leaf drying. Salinity induced significant changes in the slow part of the chlorophyll fluorescence induction curve from barley leaves.