Oxalic acid production and aluminum tolerance in Pseudomonas fluorescens

13C NMR studies on intact cells from Al-stressed Pseudomonas fluorescens incubated with citric acid or Al-citrate yielded peaks at 158 and 166 ppm that were attributable to free and complexed oxalic acid, respectively. The presence of oxalic acid was further confirmed with the aid of oxalate oxidase. These peaks were not discernable in experiments performed with cells taken from control cultures. Enzymatic analyses of cell fractions showed the highest production of oxalic acid in the inner membrane fraction of Al-stressed cells incubated with glyoxylate. There was an eight-fold increase in the synthesis of oxalic acid in the inner membrane fraction from the Al-stressed cells compared to the control cells. Although oxalic acid production was observed when citrate, Al-citrate and isocitrate were utilized as substrates, the inner membrane fraction did not mediate the formation of oxalic acid from glycine/pyruvate, glycolic acid, oxaloacetate or ascorbate. These data suggest that the increased oxalic acid production in response to Al stress is effected via the oxidation of glyoxylate.     

Naphthalene uptake by a Pseudomonas fluorescens isolate

The uptake of naphthalene has been investigated in the metabolizing cells of Pseudomonas fluorescens utilizing [1-14C]naphthalene. The uptake displayed an affinity constant (Kt) of 11 microM and a maximal velocity (Vmax) of 17 nmol.h-1.mg-1 cellular dry weight. Naphthalene uptake was not observed in a mutant strain, TG-5, which was unable to utilize naphthalene as a sole source of carbon for growth. Uptake was significantly inhibited (approximately 90%) by the presence of growth-inhibiting levels of either azide or 2,4-dinitrophenol and was sensitive to the presence of structural analogues of naphthalene. The intracellular levels of ATP were not significantly reduced by the presence of either azide or 2,4-dinitrophenol. The presence of alpha-naphthol was found to noncompetitively inhibit naphthalene uptake, displaying a Ki of 0.041 microM. It is concluded that the first step in the utilization of naphthalene by Pseudomonas fluorescens is its transport into the cell by a specific energy-linked transport system.     

Aluminum Elicits Exocellular Phosphatidylethanolamine Production in Pseudomonas fluorescens

Pseudomonas fluorescens ATCC 13525 was found to grow in a minimal mineral medium supplemented with millimolar amounts of aluminum, a known environmental toxicant. During the stationary phase of growth, the trivalent metal was localized in a phosphatidylethanolamine (PE)-containing residue. The concentration of PE in pellets ranged from 1.7 to 13.9 mg ml of culture(sup-1) in media supplemented with 1 to 30 mM aluminum. Although the gelatinous residue was observed during the stationary phase of growth, ultracentrifugation and dialysis experiments revealed that PE was produced from earlier stages of incubation and was associated with aluminum. A sharp diminution in the levels of PE and aluminum in the spent fluid was concomitant with the formation of the insoluble deposit. The aluminum content of the soluble cellular fraction increased during growth and reached an optimum of 1.85 mM of test metal at 45 h in cultures with 15 mM aluminum. Further incubation, however, led to a marked decrease in the cellular aluminum content, and during the stationary phase of growth, only trace amounts of the trivalent metal were detected in this fraction. When 45-h cells were incubated in fresh citrate medium, most of the intracellular aluminum was secreted in the spent fluid and citrate was rapidly consumed. Aluminum efflux was also observed in cultures in which d-glucose was substituted for citrate. However, no efflux of this trivalent metal was evident in media devoid of either citrate or d-glucose. Scanning electron microscopic studies and X-ray energy-dispersive analyses of the dialyzed supernatant aided in the visualization of nodule-like aluminum- and phosphorus-rich bodies associated with thread-like carbon-, oxygen-, and phosphorus-containing structures. Transmission electron microscopic and electron energy loss spectroscopic analyses revealed the presence of aluminum within bacteria after 45 h of incubation. Cells harvested after aluminum insolubilization did not shown aluminum inclusions. This aluminum-tolerant microbe may have potential application in bioremediation processes.     

Interactions of Plasmodium berghei-infected erythrocytes with complement

Incubation of radioactively labeled parasitized (Plasmodium berghei) erythrocytes (PE) with adherent peritoneal exudate cells in the presence of 10% (v/v) fresh mouse serum (NMS) resulted in the uptake of a proportion of radioactive material (PE). Inactivation of the added serum by heat or zymosan treatment resulted in diminished uptake of radioactivity. These results suggest that PE activated complement. Incubation of fresh NMS with PE reduced the hemolytic complement level of the serum as shown by its subsequent decreased ability to lyse antibody-coated rabbit red blood cells. No such effect was found when uninfected erythrocytes from either infected or uninfected blood were preincubated with fresh NMS. Thus, PE or PE-derived material activated complement. Addition of EGTA during incubation of fresh NMS with PE did not inhibit the decrease in complement level. This indicated that complement was activated by the alternative pathway. Complement levels decreased even when fresh NMS and PE were incubated in the presence of EDTA (which inhibits both classical and alternative pathway activation), suggesting that a complement activating factor (or a complement inhibitor) was released from the PE. However, lysis of PE after incubation with either fresh rabbit or guinea pig serum did not occur unless anti-mouse erythrocyte antibody was added. The production of a complement-activating factor by PE might explain part of the decreasing complement levels during infection and might enable the parasite to escape from a complement-mediated defense mechanism of the host.     

Interactions of Plasmodium berghei-Infected Erythrocytes with Complement1

Incubation of radioactively labeled parasitized (Plasmodium berghei) erythrocytes (PE) with adherent peritoneal exudate cells in the presence of 10% (v/v) fresh mouse serum (NMS) resulted in the uptake of a proportion of radioactive material (PE). Inactivation of the added serum by heat or zymosan treatment resulted in diminished uptake of radioactivity. These results suggest that PE activated complement. Incubation of fresh NMS with PE reduced the hemolytic complement level of the serum as shown by its subsequent decreased ability to lyse antibody-coated rabbit red blood cells. No such effect was found when uninfected erythrocytes from either infected or uninfected blood were preincubated with fresh NMS. Thus, PE or PE-derived material activated complement. Addition of EGTA during incubation of fresh NMS with PE did not inhibit the decrease in complement level. This indicated that complement was activated by the alternative pathway. Complement levels decreased even when fresh NMS and PE were incubated in the presence of EDTA (which inhibits both classical and alternative pathway activation), suggesting that a complement activating factor (or a complement inhibitor) was released from the PE. However, lysis of PE after incubation with either fresh rabbit or guinea pig serum did not occur unless anti-mouse erythrocyte antibody was added. The production of a complement-activating factor by PE might explain part of the decreasing complement levels during infection and might enable the parasite to escape from a complement-mediated defense mechanism of the host.     

Organic acid exudation induced by phosphorus deficiency and/or aluminium toxicity in two contrasting soybean genotypes

Experiments in nutrient solution were conducted to investigate the exudation of organic acids (OAs) induced by phosphorus deficiency (–P) and/or aluminium toxicity (+Al) in two contrasting soybean genotypes as related to internal OA concentration and related enzyme activities. Baxi 10 (BX10), a known P‐efficient soybean (Glycine max[L] Merr.) genotype, was shown to be more resistant to +Al than a P‐inefficient genotype Bendi 2 (BD2), indicating the potential of selecting soybean cultivars with dual resistance to –P and +Al. The two contrasting genotypes were further characterized for root exudation and formation of oxalate, malate and citrate and their related enzyme activities in response to –P, +Al or both combined. –P significantly induced malate and oxalate exudation from both soybean genotypes, although the P‐efficient BX10 tended to excrete much more oxalate than the P‐inefficient BD2. The +Al treatment triggered citrate efflux from both genotypes, with BX10 having a much greater efflux rate than BD2. Interestingly, –P did not appear to induce citrate exudation, whereas +Al had no obvious effect on malate or oxalate exudation from the two genotypes. The exudation of OAs was generally diminished under the coupled stress of –P and +Al in comparison with either single stress, implying a possible antagonistic effect of the two stresses on OA exudation. Root malate content was negatively correlated with its exudation in BX10 but positively in BD2. A similar tendency was observed for oxalate content and exudation only with less magnitude. Determination of six related enzymes, phosphoenolpyruvate carboxylase (PEPC), phosphoenolpyruvate phosphatase (PEPP), malate enzyme (ME), isocitrate dehydrogenase (ICDH), malate dehydrogenase (MDH), and pyruvate kinase (PK), in the root tips showed that their activities were not significantly altered during the early stage of treatments (2 and 4 days) whereas at 14 days after stress imposition, the activities of PEPC, PEPP, ME and ICDH were generally enhanced for both genotypes. However, the activity of these enzymes did not appear to be correlated with OA exudation or formation. This study clearly demonstrates that OA exudation is differentially induced by –P and +Al in soybean plants, with specific induction of oxalate and malate by –P and citrate by +Al. The lack of a close relationship between OA exudation and internal concentration or enzyme activities may suggest that the regulation of OA formation and exudation by –P and/or +Al could be imposed at different stages.     

Modulation of TCA cycle enzymes and aluminum stress in Pseudomonas fluorescens.

Oxalic acid plays a pivotal role in the adaptation of the soil microbe Pseudomonas fluorescens to aluminum (Al) stress. Its production via the oxidation of glyoxylate necessitates a major reconfiguration of the enzymatic reactions involved in the tricarboxylic acid (TCA) cycle. The demand for glyoxylate, the precursor of oxalic acid appears to enhance the activity of isocitrate lyase (ICL). The activity of ICL, an enzyme that participates in the cleavage of isocitrate to glyoxylate and succinate incurred a 4-fold increase in the Al-stressed cells. However, the activity of isocitrate dehydrogenase, a competitor for the substrate isocitrate, appeared to be diminished in cells exposed to Al compared to the control cells. While the demand for oxalate in Al-stressed cells also negatively influenced the activity of the enzyme alpha-ketoglutarate dehydrogenase complex, no apparent change in the activity of malate synthase was recorded. Thus, it appears that the TCA cycle is tailored in order to generate the necessary precursor for oxalate synthesis as a consequence of Al-stress.     

Changes and role of adrenoceptors in PC12 cells after phenylephrine administration and apoptosis induction

The present study addresses the hypothesis that adrenergic regulation modulates the effect of apoptosis. Therefore we studied, whether α1-adrenergic receptor's agonist phenylephrine (PE) can affect or induce apoptosis in rat pheochromocytoma (PC12) cells. We have shown that PE treatment did not increase level of the apoptosis, or level of the caspase 3 mRNA. When apoptosis was induced in the presence of PE, caspase 3 mRNA was significantly increased, while the percentage of apoptotic cells remained unchanged compared to apoptotic group without PE. During this process, α1D-, β2- and β3-adrenergic receptors (ARs) were upregulated. Since all these three types of ARs are differently localized in the cell, we assume that mutual communication of all three ARs is crucial to participate in this signaling and during development of apoptosis, some of these systems might translocate. Another important system in handling noradrenaline during apoptosis might be noradrenaline transporter (NET), since it was downregulated in apoptotic cells treated with PE, compared to untreated apoptotic cells. However, precise mechanism of mutual communication among all these systems remains to be elucidated.     

A model study of factors involved in adhesion of Pseudomonas fluorescens to meat.

A study was undertaken to investigate the factors involved in the adhesion of Pseudomonas fluorescens to model meat surfaces (tendon slices). Adhesion was fast (less than 2.5 min) and was not suppressed by killing the cells with UV, gamma rays, or heat, indicating that physiological activity was not required. In various salt solutions (NaCl, KCl, CaCl2, MgCl2), adhesion increased with increasing ionic strength up to 10 to 100 mM, suggesting that, at low ionic strengths, electrostatic interactions were involved in the adhesion process. At higher ionic strengths (greater than 10 to 100 mM) or in the presence of Al3+ ions, adhesion was sharply reduced. Selectively blocking of carboxyl or amino groups at the cell surface by chemical means did not affect adhesion. These groups are therefore not directly involved in an adhesive bond with tendon. Given a sufficient cell concentration (10(10) CFU.ml-1) in the adhesion medium, the surface of tendon was almost entirely covered with adherent bacteria. This suggests that if the adhesion is specific, the attachment sites on the tendon surface must be located within collagen or proteoglycan molecules.     

A model study of factors involved in adhesion of Pseudomonas fluorescens to meat.

A study was undertaken to investigate the factors involved in the adhesion of Pseudomonas fluorescens to model meat surfaces (tendon slices). Adhesion was fast (less than 2.5 min) and was not suppressed by killing the cells with UV, gamma rays, or heat, indicating that physiological activity was not required. In various salt solutions (NaCl, KCl, CaCl2, MgCl2), adhesion increased with increasing ionic strength up to 10 to 100 mM, suggesting that, at low ionic strengths, electrostatic interactions were involved in the adhesion process. At higher ionic strengths (greater than 10 to 100 mM) or in the presence of Al3+ ions, adhesion was sharply reduced. Selectively blocking of carboxyl or amino groups at the cell surface by chemical means did not affect adhesion. These groups are therefore not directly involved in an adhesive bond with tendon. Given a sufficient cell concentration (10(10) CFU.ml-1) in the adhesion medium, the surface of tendon was almost entirely covered with adherent bacteria. This suggests that if the adhesion is specific, the attachment sites on the tendon surface must be located within collagen or proteoglycan molecules.     

Form of Al changes with Al concentration in leaves of buckwheat

Buckwheat (Fagopyrum esculentum Moench. cv. Jianxi) is known as an Al-accumulating plant. The process leading to the accumulation of Al in the leaves was investigated, focusing on the chemical form of Al using 27Al-nuclear magnetic resonance. Leaves with different Al concentrations were prepared by growing buckwheat on a very acidic soil (Andosol) amended with or without CaCO3 (1 or 3 g x kg-1 soil). When the Al concentration of the leaves was lower, only one major signal was observed at a chemical shift of 16.1 ppm, which was assigned to an Al-oxalate complex at a 1:3 ratio. However, when the Al concentration of the leaves increased to a high level (e.g. 12 g Al kg-1), an additional signal at a chemical shift of 11.2 ppm was observed. This signal was assigned to an Al-citrate complex at a 1:1 ratio. In the leaf with a high Al concentration, both Al-oxalate (1:3) and Al-citrate (1:1) were detected in marginal and middle parts, while only Al-oxalate was detected in the basal part. The oxalate concentration did not differ very much between leaves with low and high Al concentrations at the same position, while citrate concentration significantly increased with increasing Al concentration when the oxalate/Al ratio became lower than 3.0. As the Al-citrate complex has been demonstrated to be the form of transport in the xylem, the results suggest that when internal oxalate is enough to form a complex with Al at a 3:1 ratio in the leaves with a low Al concentration, Al-citrate converts to Al-oxalate. However, this conversion does not occur in the leaves with a very high Al concentration, resulting in the coexistence of both Al-oxalate and Al-citrate complexes.     

Comparative studies on the effect of a protein-synthesis inhibitor on aluminium-induced secretion of organic acids from Fagopyrum esculentum Moench and Cassia tora L. roots

Aluminium (Al)-induced secretion of organic acids from plant roots is considered a mechanism of Al resistance, but the processes leading to the secretion of organic acids are still unknown. In the present study, a protein-synthesis inhibitor, cycloheximide (CHM), was used to investigate its effect on Al-induced organic acid secretion in a pattern I (rapid exudation of organic acids under Al stress) plant buckwheat (Fagopyrum esculentum Moench) and a pattern II (exudation of organic acids was delayed by several hours under Al stress) plant Cassia tora L. A dose-response experiment showed that the secretion of oxalate by buckwheat roots was not affected by CHM when added in the range from 0 to 50 microM, with or without exposure to 100 microm Al, but the secretion of citrate was completely inhibited by 30 microM CHM in C. tora. A time-course experiment showed that even prolonged exposure to 20 microM CHM did not affect oxalate secretion in buckwheat, but significantly inhibited citrate secretion in C. tora. However, citrate synthase (CS) activity in C. tora was not affected during 12 h exposure to 100 microM Al when compared with that in control roots, although CHM can inhibit CS activity effectively. These results indicated that CS activity was not related to Al-regulated citrate efflux in C. tora. The total protein was decreased by 14.0% and 32.3% in C. tora and buckwheat root tip, respectively, after 3-h treatment with 20 microM CHM. A 3-h pulse with 20 microM CHM completely inhibited citrate efflux in C. tora during the next 6-h exposure to Al, although a small amount of citrate was exuded after 9-h exposure. However, oxalate efflux in buckwheat was not influenced by a similar treatment. In buckwheat, a 3-h pulse with 100 microM Al maintained oxalate secretion at a high level during the next 9 h, with or without CHM treatment. Conversely, in C. tora a 6-h pulse with 100 microM Al induced significant secretion of citrate which was inhibited by the CHM. Taken together, these findings suggest that both de novo synthesis and activation of an anion channel are needed for Al-induced secretion of citrate in C. tora, but in buckwheat the plasma membrane protein responsible for oxalate secretion pre-exists.     

Oxalate transport via the sulfate/HCO3 exchanger in rabbit renal basolateral membrane vesicles

We evaluated the mechanism of oxalate transport in basolateral membrane vesicles isolated from the rabbit renal cortex. An outward HCO3- gradient induced the transient uphill accumulation of oxalate and sulfate, indicating the presence of oxalate/HCO3- exchange and sulfate/HCO3- exchange. For oxalate, sulfate, or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, the K1/2 value for oxalate/HCO3- exchange was nearly identical to that for sulfate/HCO3- exchange, suggesting that both exchange processes occur via the same transport system. This was further supported by the finding of sulfate/oxalate exchange. Thiosulfate/sulfate exchange and thiosulfate/oxalate exchange were also demonstrated, but a variety of other tested anions including Cl-, p-aminohippurate, and lactate did not exchange for sulfate or oxalate. Na+ did not affect sulfate or oxalate transport, indicating that neither anion undergoes Na+ co-transport or Na+-dependent anion exchange in these membrane vesicles. Finally, we found that the stoichiometry of exchange is 1 sulfate or oxalate per 2 HCO3-, or a thermodynamically equivalent process. We conclude that oxalate, but not other organic or inorganic anions of physiologic importance, can share the sulfate/HCO3- exchanger in renal basolateral membrane vesicles. In series with luminal membrane oxalate/Cl- (formate) exchange, exchange of oxalate for HCO3- or sulfate across the basolateral membrane provides a possible transcellular route for oxalate transport in the proximal tubule.     

Receptor-mediated internalization of Pseudomonas toxin by mouse fibroblasts

Pseudomonas exotoxin (PE) was used as a probe to study the mechanism by which protein ligands are internalized by mammalian cells. Both biochemical and electron microscopic methods were used to look at the internalization of PE by mouse LM cell fibroblasts. Our data suggest that PE enters cells by receptor-mediated endocytosis, a process previously thought to be restricted to the entry of biologically significant molecules such as lysosomal enzymes and peptide hormones. Biochemical studies showed that methylamine (20 mM) and chloroquine (10 microM) protected LM cells from the action of PE. Full protection was observed if methylamine or chloroquine was added to the monolayers simultaneously with toxin or if they were added up to 10 min after toxin binding. Later addition of amine or chloroquine afforded partial protection to the monolayers. With immunoelectron microscopy we observed that in the cold toxin bound diffusely to the cell surface but was rapidly internalized when cells were warmed to 37 degrees C. In the presence of methylamine, chloroquine or ammonium chloride, internalization did not occur. We propose that PE enters mouse fibroblasts by receptor-mediated endocytosis and that chloroquine and methylamine, agents which are known to block this process, prevent expression of toxicity.     

Oxalate:formate exchange. The basis for energy coupling in Oxalobacter

In the Gram-negative anaerobe, Oxalobacter formigenes, the generation of metabolic energy depends on the transport and decarboxylation of oxalate. We have now used assays of reconstitution to study the movements of oxalate and to characterize the exchange of oxalate with formate, its immediate metabolic derivative. Membranes of O. formigenes were solubilized with octyl-beta-D-glucopyranoside in the presence of 20% glycerol and Escherichia coli phospholipid, and detergent extracts were reconstituted by detergent dilution. [14C]Oxalate was taken up by proteoliposomes loaded with unlabeled oxalate, but not by similarly loaded liposomes or by proteoliposomes containing sulfate in place of oxalate. Oxalate transport did not depend on the presence of sodium or potassium, nor was it affected by valinomycin (1 microM), nigericin (1 microM), or a proton conductor, carbonylcyanide-p-trifluoromethoxyphenylhydrazone (5 microM) when potassium was at equal concentration on either side of the membrane. Such data suggest the presence of an overall neutral oxalate self-exchange, independent of common cations or anions. Kinetic analysis of the reaction in proteoliposomes gave a Michaelis constant (Kt) for oxalate transport of 0.24 mM and a maximal velocity (Vmax) of 99 mumol/min/mg of protein. A direct exchange of oxalate and formate was indicated by the observations that formate inhibited oxalate transport and that delayed addition of formate released [14C]oxalate accumulated during oxalate exchange. Moreover, [14C]formate was taken up by oxalate-loaded proteoliposomes (but not liposomes), and this heterologous reaction could be blocked by external oxalate. Further studies, using formate-loaded proteoliposomes, suggested that the heterologous exchange was electrogenic. Thus, for assays in which N-methylglucamine served as both internal and external cation, formate-loaded particles took up oxalate at a rate of 2.4 mumol/min/mg of protein. When external or internal N-methylglucamine was replaced by potassium in the presence of valinomycin, there was, respectively, a 7-fold stimulation or an 8-fold inhibition of oxalate accumulation, demonstrating that net negative charge moved in parallel with oxalate during the heterologous exchange. The work summarized here suggests the presence of an unusually rapid and electrogenic oxalate2-:formate1- antiport in membranes of O. formigenes. Since a proton is consumed during the intracellular decarboxylation that converts oxalate into formate plus CO2, antiport of oxalate and formate would play a central role in a biochemical cycle consisting of (a) oxalate influx, (b) oxalate decarboxylation, and (c) formate efflux.(ABSTRACT TRUNCATED AT 400 WORDS)     

Protein kinase C modulators influence meiosis kinetics but not fertilizability of mouse oocytes.

The role of protein kinase C (PKC) in the successive steps of mouse oocyte meiotic process was investigated. We have used either OAG, an analog of diacylglycerol, or mezerein, a nonphorbol ester diterpene, less tumor promoting than phorbol esters, as PKC activators, and staurosporine as PKC inhibitor. Cumulus-free oocytes were cultured in minimum essential medium with each of these PKC modulators and maturation stages were screened every two hours until the end of the process. Both PKC activators prevented GVBD at each tested dose for 4 hr (OAG) and 8 hr (mezerein), and decreased the frequencies of PB oocytes. The inhibitory effects of both activators were dose dependent and reversible. The addition of OAG to the culture medium after GVBD occurrence (i.e., after 4 hrs) did not affect PB extrusion whereas similar addition of mezerein significantly decreased the frequency of PB oocytes. Inhibition of PKC by staurosporine accelerated GVBD and increased the frequency of PB extrusion. When staurosporine was added after GVBD, PB extrusion occurred earlier but PB oocyte frequency was not increased. Fertilizability was not affected when oocyte maturation occurred in the presence of any of these substances despite the delay in maturation process. These results clearly indicate that the PKC pathway is involved in mouse oocyte meiotic process: activation of the enzyme would arrest meiotic process whereas its inhibition would participate in meiosis induction.     

Alanine Efflux across the Serosal Border of Turtle Intestine

The exit of alanine across the serosal border of the epithelial cells of turtle intestine was measured by direct and indirect techniques. A decrease or an increase in cell Na did not affect the amino acid flux from cell to serosal solution. Cells loaded with Na and alanine did not exhibit any extrusion of alanine when their serosal membranes were exposed to an Na-free medium containing alanine. However, substantial amino acid extrusion was observed across the mucosal cell border under similar conditions. Although alanine flux across the serosal membrane appeared to be Na-independent, it showed a tendency toward saturation as cellular alanine concentration was elevated. The results are consistent with the postulate that the serosal and mucosal membranes of intestinal cells are asymmetrical with respect to amino acid transport mechanisms. The serosal membrane appears to have an Na-independent carrier-mediated mechanism responsible for alanine transport while transport across the mucosal border involves an Na-dependent process.     

Tetradecylthioacetic acid (a 3-thia fatty acid) impairs secretion of oleic acid-induced triacylglycerol-rich lipoproteins in CaCo-2 cells

The fatty acid analogue tetradecylthioacetic acid (TTA) has previously been shown to decrease triacylglycerol secretion in CaCo-2 cells (Gedde-Dahl et al., J. Lipid Res. 36 (1995) 535-543). The present study was designed to further elucidate the effect of TTA on lipoprotein production in CaCo-2 cells. TTA did not affect oleic acid-induced triacylglycerol synthesis, but it significantly decreased secretion of newly synthesized triacylglycerol when compared to cells incubated with oleic acid alone or oleic acid in combination with palmitic acid. In contrast, pulse-chase experiments showed no difference in the amount of labeled triacylglycerol secreted from cells exposed to either fatty acid combination during the chase period, indicating that TTA did not affect the secretory process in general. Cells incubated with TTA alone secreted triacylglycerol present at 1.025相似文献   

Al-induced efflux of organic acid anions is poorly associated with internal organic acid metabolism in triticale roots

The secretion of organic acid anions from roots has been identified as a mechanism of resistance to Al. However, the process leading to the secretion of organic acid anions is poorly understood. The effect of Al on organic acid metabolism was investigated in two lines of triticale (xTriticosecale Wittmark) differing in Al-induced secretion of malate and citrate and in Al resistance. The site of Al-induced secretion of citrate and malate from a resistant line was localized to the root apices (terminal 5 mm). The levels of citrate (root apices and mature root segments) and malate (mature segments only) in roots increased during exposure to Al, but similar changes were observed in both triticale genotypes. The in vitro activities of four enzymes involved in malate and citrate metabolism (citrate synthase, phosphoenolpyruvate carboxylase, malate dehydrogenase, and NADP-isocitrate dehydrogenase) were similar for sensitive and resistant lines in both root apices and mature root segments. The response of these enzymes to pH did not differ between tolerant and sensitive lines or in the presence and absence of Al. Moreover, cytoplasmic and vacuolar pH were not affected by exposure to Al in either line. Together, these results indicate that the Al-dependent efflux of organic acid anions from the roots of triticale is not regulated by their internal levels in the roots or by the capacity of the root cells to synthesize malate and citrate.