摘要:

Na+/H+exchange activity in barley (Hordeum vulgarecv CM-72) root tonoplast was induced by Na+even in the presence of inhibitors of protein synthesis. Induction occurred with a half-time of only 15 minutes. When salt-treated roots were transferred to a nutrient solution containing no Na+, the activity disappeared with a similar time course. The data suggest that Na+/H+exchange was due to activation of an existing protein rather than tode novoprotein synthesis.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

Bacterial exopolysaccharide (EPS) was extracted from infected leaves of several host plants inoculated with phytopathogenic strains ofPseudomonas syringaepathovars. Extraction was by a facilitated diffusion procedure or by collection of intercellular fluid using a centrifugation method. The extracted EPS was purified and characterized. All bacterial pathogens which induced watersoaked lesions on their host leaves, a characteristic of most members of this bacterial group, were found to produce alginic acid (a polymer consisting of varying ratios of mannuronic and guluronic acids). Only trace amounts of bacterial EPS could be isolated from leaves inoculated with a pathovar (pv.syringae) which does not induce the formation of lesions with a watersoaked appearance. Guluronic acid was either present in very low amounts or absent in the alginic acid preparations. All bacterial alginates were acetylated (7-11%). Levan (a fructan) was apparently not produced as an EPSin vivoby any of the pathogens tested.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

Key branchpoint enzymes of aromatic amino acid biosynthesis, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DS) and chorismate mutase (CM), have previously been shown to exist as separate compartmentalized isozymes in the chloroplasts and cytosol of tobacco, sorghum and spinach. Although additional examples of plants containing these isozyme pairs are accumulating, some studies in the literature report the presence of only the single plastidic DS or CM enzyme. Such apparent exceptions contradict the universality of pathway organization existing in higher plants that is implied by the dual-pathway hypothesis of aromatic biosynthesis. Since potato (Solanum tuberosum) exemplifies a case where only a single species of both DS and CM have been reported, we selected this system for further analysis. The DS-Mn and DS-Co isozyme pair, exhibiting all of the differential properties described inNicotiana silvestris, have now been identified inS. tuberosum.Likewise, partial purification via DEAE-cellulose chromatography revealed two isozymes of CM in disks excised from tubers ofS. tuberosum.The differential regulatory properties of these isozymes were comparable to the CM-1 and CM-2 isozymes ofN. silvestris.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

To study the import of polypeptide precursors we have adapted and compared two procedures for the isolation of competent chloroplasts from the green unicellular alga,Chlamydomonas reinhardtii: silicasol gradient centrifugation and elutriation. The chloroplasts actively import the precursor of the small subunit of ribulose bisphosphate carboxylase-oxygenasein vitro.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

For the first time, the31P nuclear magnetic resonance technique has been used to study the properties of isolated vacuoles of plant cells, namely the vacuolar pH and the inorganic phosphate content.Catharanthus roseuscells incubated for 15 hours on a culture medium enriched with 10 millimolar inorganic phosphate accumulated large amounts of inorganic phosphate in their vacuoles. Vacuolar phosphate ions were largely retained in the vacuoles when protoplasts were prepared from the cells and vacuoles isolated from the protoplasts. Vacuolar inorganic phosphate concentrations up to 150 millimolar were routinely obtained. Suspensions prepared with 2 to 3×106vacuoles per milliliter from the enrichedC. roseuscells have an internal pH value of 5.50±0.06 and a meantrans-tonoplastΔpH of 1.56±0.07. Reliable determinations of vacuolar and external pH could be made by using accumulation times as low as 2 minutes. These conditions are suitable to follow the kinetics of H+exchanges at the tonoplast. The31P nuclear magnetic resonance technique also offered the possibility of monitoring simultaneously the stability of thetrans-tonoplast pH and phosphate gradients. Both appeared to be reasonably stable over several hours. The buffering capacity of the vacuolar sap around pH 5.5 has been estimated by several procedures to be 36±2 microequivalents per milliliter per pH unit. The increase of the buffering capacity due to the accumulation of phosphate in the vacuoles is, in large part, compensated by a decrease of the intravacuolar malate con

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

The vacuolar pH and thetrans-tonoplastΔpH modifications induced by the activity of the two proton pumps H+-ATPase and H+-PPase and by the proton exchanges catalyzed by the Na+/H+and Ca2+/H+antiports at the tonoplast of isolated intact vacuoles prepared fromCatharanthus roseuscells enriched in inorganic phosphate (Y Mathieu et al 1988 Plant Physiol [in press]) were measured using the31P NMR technique. The H+-ATPase induced an intravacuolar acidification as large as 0.8 pH unit, building atrans-tonoplastΔpH up to 2.2 pH units. The hydrolysis of the phosphorylated substrate and the vacuolar acidification were monitored simultaneously to estimate kinetically the apparent stoichiometry between the vectorial proton pumping and the hydrolytic activity of the H+-ATPase. A ratio of H+translocated/ATP hydrolyzed of 1.97±0.06 (mean±standard error) was calculated. Pyrophosphate-treated vacuoles were also acidified to a significant extent. The H+-PPase at 2 millimolar PPi displayed hydrolytic and vectorial activities comparable to those of the H+-ATPase, building a steady stateΔpH of 2.1 pH units. Vacuoles incubated in the presence of 10 millimolar Na+were alkalinized by 0.4 to 0.8 pH unit. It has been shown by using23Na NMR that sodium uptake was coupled to the H+efflux and occurred against rather large concentration gradients. For the first time, the activity of the Ca2+/H+antiport has been measured on isolated intact vacuoles. Ca2+uptake was strongly inhibited by NH4Cl or gramicidin. Vacuoles incubated with 1 millimolar Ca2+were alkalinized by about 0.6 pH unit and this H+efflux was associated to a Ca2+uptake as demonstrated by measuring the external Ca2+concentration with a calcium specific electrode. Steady state accumulation ratios of Ca2+as high as 100 were reached for steady state external concentrations about 200 micromolar. The rate of Ca2+uptake appeared markedly amplified in intact vacuoles when compared to tonoplast vesicles but the antiport displayed a much lower affinity for calcium. The different behavior of intact vacuoles compared to vesicles appears mainly to be due to differences in the surface to volume ratio and in the rates of dissipation of the pH gradient. Despite its low affinity, the Ca2+/H+antiport has a high potential capacity to regulate cytoplasmic concentration of cal

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

In high inorganic carbon grown (1% CO2[volume/volume]) cells of the cyanobacteriumSynechococcusPCC7942, the carbonic anhydrase (CA) inhibitor, ethoxyzolamide (EZ), was found to inhibit the rate of CO2uptake and to reduce the final internal inorganic carbon (Ci) pool size reached. The relationship between CO2fixation rate and internal Ciconcentration in high Cigrown cells was little affected by EZ. This suggests that in intact cells internal CA activity was unaffected by EZ. High Cigrown cells readily took up CO2but had little or no capacity for HCO3−uptake. These cells appear to possess a CO2utilizing Cipump that has a CA-like function associated with the transport step such that HCO3−is the species delivered to the cell interior. This CA-like step may be the site of inhibition by EZ. Low Cigrown cells possess both CO2uptake and HCO3−uptake activities and EZ inhibited both activities to a similar degree, suggesting that a common step in CO2and HCO3−uptake (such as the Cipump) may have been affected. The inhibitor had no apparent effect on internal CO2/HCO3−equilibria (internal CA function) in low Cigr

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

Cells of the cyanobacterium,SynechococcusPCC7942, grown under high inorganic carbon (Ci) conditions (1% CO2; pH 8) were found to be photosynthetically dependent on exogenous CO2. This was judged by the fact that they had a similar photosynthetic affinity for CO2(K0.5[CO2] of 3.4-5.4 micromolar) over the pH range 7 to 9 and that the low photosynthetic affinity for Cimeasured in dense cell suspensions was improved by the addition of exogenous carbonic anhydrase (CA). The CA inhibitor, ethoxyzolamide (EZ), was shown to reduce photosynthetic affinity for CO2in high Cicells. The addition of 200 micromolar EZ to high Cicells increasedK0.5(CO2) from 4.6 micromolar to more than 155 micromolar at pH 8.0, whereas low Cicells (grown at 30 microliters CO2per liter of air) were less sensitive to EZ. EZ inhibition in high and low Cicells was largely relieved by increasing exogenous Ciup to 100 millimolar. Lipid soluble CA inhibitors such as EZ and chlorazolamide were shown to be the most effective inhibitors of CO2usage, whereas water soluble CA inhibitors such as methazolamide and acetazolamide had little or no effect. EZ was found to cause a small drop in photosystem II activity, but this level of inhibition was not sufficient to explain the large effect that EZ had on CO2usage. High Cicells ofAnabaena variabilisM3 andSynechocystisPCC6803 were also found to be sensitive to 200 micromolar EZ. We discuss the possibility that the inhibitory effect of EZ on CO2usage in high Cicells ofSynechococcusPCC7942 may be due to inhibition of a `CA-like' function associated with the CO2utilizing Cipump or due to inhibition of an internal CA activity, thus affecting CO2supply to ribulose bisphosphate carboxylase-oxygenase.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

Intact cells and crude homogenates of high (1% CO2) and low dissolved inorganic carbon (Ci) (30-50 microliters per liter of CO2) grownSynechococcusPCC7942 have carbonic anhydrase (CA)-like activity, which enables them to catalyze the exchange of18O from CO2to H2O. This activity was studied using a mass spectrometer coupled to a cuvette with a membrane inlet system. Intact high and low Cicells were found to contain CA activity, separated from the medium by a membrane which is preferentially permeable to CO2. This activity is most apparent in the light, where18O-labeled CO2species are being taken up by the cells but the effluxing CO2has lost most of its label to water. In the dark, low Cicells catalyze the depletion of the18O enrichment of CO2and this activity is inhibited by both ethoxyzolamide and 2-(trifluoromethoxy)carbonyl cyanide. This may occur via a common inhibition of the Cipump and the Cipump is proposed as a potential site for the exchange of18O. CA activity was measurable in homogenates of both cell types but was 5- to 10-fold higher in low Cicells. This was inhibited by ethoxyzolamide with an I50of 50 to 100 micromolar in both low and high Cicells. A large proportion of the internal CA activity appears to be pelletable in nature. This pelletability is increased by the presence of Mg2+in a manner similar to that of ribulose bisphosphate carboxylase-oxygenase activity and chlorophyll (thylakoids) and may be the result of nonspecific aggregation. Separation of crude homogenates on sucrose gradients is consistent with the notion that CA and ribulose bisphosphate carboxylase-oxygenase activity may be associated with the same pelletable fraction. However, we cannot unequivocally establish that CA is located within the carboxysome. The sucrose gradients show the presence of separate soluble and pelletable CA activity. This may be due to the presence of separate forms of the enzyme or may arise from the same pelletable association which is unstable during extraction.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB

摘要:

Photosynthesis and light O2-uptake of the aerial portion of the CAM plantAnanas comosus(L.) merr. were studied by CO2and O2gas exchange measurements. The amount of CO2which was fixed during a complete day-night cycle was equal to the amount of total net O2evolved. This finding justifies the assumption that in each time interval of the light period, the difference between the rates of net O2-evolution and of net light atmospheric CO2-uptake give the rates of malate-decarboxylation-dependent CO2assimilation. Based upon this hypothesis, the following photosynthetic characteristics were observed: (a) From the onset of the light to midphase IV of CAM, the photosynthetic quotient (net O2evolved/net CO2fixed) was higher than 1. This indicates that malate-decarboxylation supplied CO2for the photosynthetic carbon reduction cycle during this period. (b) In phase III and early phase IV, the rate of CO2assimilation deduced from net O2-evolution was 3 times higher than the maximum rate of atmospheric CO2-fixation during phase IV. A conceivable explanation for this stimulation of photosynthesis is that the intracellular CO2-concentration was high because of malate decarboxylation. (c) During the final hours of the light period, the photosynthetic quotient decreased below 1. This may be the result of CO2-fixation by phosphoenolpyruvate-carboxylase activity and malate accumulation. Based upon this hypothesis, the gas exchange data indicates that at least 50% of the CO2fixed during the last hour of the light period was stored as malate. Light O2-uptake determined with18O2showed two remarkable characteristics: from the onset of the light until midphase IV the rate of O2-uptake increased progressively; during the following part of the light period, the rate of O2-uptake was 3.5 times higher than the maximum rate of CO2-uptake. When malate decarboxylation was reduced or suppressed after a night in a CO2-free atmosphere or in continuous illumination, the rate of O2-uptake was higher than in the control. This supports the hypothesis that the low rate of O2-uptake in the first part of the light period is due to the inhibition of photorespiration by increased intracellular CO2concentration because of malate decarboxylation. In view of the law of gas diffusion and the kinetic properties of the ribulose-1,5-bisphosphate carboxylase/oxygenase, O2and CO2gas exchange suggest that at the end of the light period the intracellular CO2concentration was very low. We propose that the high ratio of O2-uptake/CO2-fixation is principally caused by the stimulation of photorespiration during this period.

ISSN:0032-0889    

出版商:American Society of Plant Biologists    

年代:1989

数据来源: ASPB