Journal of Experimental Botany, Vol. 52, No. 359, pp. 1315-1322,
June 1, 2001
© 2001 Oxford University Press
Original Papers |
Uptake of glycine by non-mycorrhizal Lolium perenne
The Macaulay Land Use Research Institute, Craigiebuckler, Aberdeen AB15 8QH, Scotland, UK
Received 20 September 2000; Accepted 22 January 2001
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Abstract |
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Plants of Lolium perenne were grown in sterile solution culture. 15N-labelled glycine (Gly) coupled with gas chromatograph mass spectrometry was used to prove that non-mycorrhizal plants of L. perenne are capable of acquiring N in the form of intact Gly. It was estimated that a minimum of 80% of Gly-N uptake, over a 3 h period, was as intact Gly, though possible processes resulting in deviation from this estimate are discussed. The relative incorporation of 15N derived from Gly uptake into serine (Ser) compared with other amino acids in the root amino acid pool suggested the enzyme serine:glyoxylate aminotransferase was at least partly responsible for the synthesis of Ser from Gly. Defoliation was shown to reduce Gly uptake by L. perenne. The addition of either 25 mol m-3 sucrose or 50 mol m-3 glucose to the uptake solution of defoliated plants increased Gly-N uptake compared with both defoliated plants without sugars and with undefoliated plants. Addition of a glucose analogue, 3-O-methyl-D-glucopyranose, that is absorbed but not metabolized by plants, did not affect Gly uptake by defoliated plants. Increasing pH from 3.5 to 9.2 caused a reduction in Gly uptake. Results of the effects of defoliation and pH are consistent with Gly uptake by L. perenne being by an energy-dependent proton symport. When either
or Gly were supplied to plants at equimolar concentrations, uptake was five times greater than that of Gly at pH 6 and 13 times greater at pH 9. Key words: Lolium perenne, glycine, ammonium, uptake, defoliation.
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Introduction |
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Contrasting results have been reported in the degree to which uptake of amino acid N by grasses occurred as intact amino acids. Natural abundance measurements of 15N have been used to suggest that plants differing in mycorrhizal associations and growing in nutrient-limited environments, access different sources of soil N (Michelsen et al., 1996
, 1998). In heath or forest tundra ecosystems, non-mycorrhizal graminoids relied mainly on acquisition of inorganic N, whilst plants associated with ericoid or ectomycorrhizal fungi were able to access organic N sources to a greater extent (Michelsen et al., 1996, 1998).
The acquisition of N by Lolium perenne from patches of 15N/13C dual labelled lysine (Lys) (13C labelled at position 1-C) was investigated (Hodge et al., 1999a, b). Although 15N uptake by the plants was rapid, no increase in plant 13C content was observed (Hodge et al., 1999a, b), suggesting the Lys was metabolized outside the plant prior to 15N uptake. In contrast, Näsholm and co-workers using 15N/13C dual labelled glycine (Gly) (13C universally labelled), estimated that a minimum of 19% of the Gly uptake by Phleum pratense growing in agricultural soil was as the intact amino acid (Näsholm et al., 2000). This figure was increased to 64% for plants of Deschampsia flexuosa growing in a boreal forest (Näsholm et al., 1998). Using similar dual labelling techniques, discrepancy in the degree to which Gly was estimated to be absorbed as an intact molecule by sedges (Schimel and Chapin, 1996; Lipson and Monson, 1998) was considered by Lipson and Monson to be due to differences in the position of the 13C label. They suggested the carboxyl carbon of Gly might be more readily respired than the methylene carbon, resulting in an underestimation of N uptake in the form of intact Gly.
Nitrogen-15 labelled amino acids coupled with gas chromatograph mass spectrometry (GC-MS) was used previously to show that for Lolium perenne growing on a clay substrate, both non-mycorrhizal plants and plants infected with the AM fungus Glomus fasciculatum took up a large proportion of serine (Ser) as the intact amino acid (Cliquet et al., 1997). Uptake of Ser-N was greater in AM-infected than non-mycorrhizal plants. Evidence for direct uptake of aspartic acid (Asp) by equivalent plants was less clear (Cliquet et al., 1997). Although the clay substrate used was originally sterilized, no attempt was made to maintain sterility over a 49 d growth period, and they considered that a greater microbial breakdown of Asp compared with Ser in the clay could not be discounted (Cliquet et al., 1997). In non-sterile systems, plants will compete with microbes for amino acids (Schimel and Chapin, 1996; Lipson and Monson, 1998; Hodge et al., 1999b). The competitive ability of plants compared with microbes for amino acids has been shown to be amino acid specific (Lipson et al., 1999). In soil, plants also compete for amino acids against chemical adsorption onto solid phases, again such competition being amino acid specific (Jones and Hodge, 1999).
Gly is a common amino acid in the soil solution of many ecosystems (Kielland, 1995; Schmidt and Stewart, 1999; Streeter et al., 2000). It has the potential to be a source of N to plants in grazed pastures as it is returned to the soil in urine (Bathurst, 1952; Bristow et al., 1992). However, the rapid degradation of urea to ammonium following urine deposition on soil (Thomas et al., 1988) means is also likely to be a source of N for plants at this time. This study investigates the uptake of Gly by a grass species commonly found in pastures, Lolium perenne. Although plants compete well against microbes for Gly compared with other amino acids (Lipson et al., 1999), plants were grown in solution culture under sterile conditions avoiding plant competition for Gly both with microbes and chemical adsorption processes. 15N labelled Gly coupled with GC-MS was used to (1) establish if non-mycorrihzal plants of Lolium perenne are capable of Gly-N uptake and if so determine (2) whether Gly-N is taken up as an intact Gly entity and (3) how uptake of Gly-N is influenced by defoliation and changes in pH. The pH response of Gly uptake was compared with that of supplied at an equimolar concentration of N.
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Materials and methods |
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Growth of plant material
Seeds of Lolium perenne L. cv. Parcour were surface-sterilized in 0.5% (v/v) peracetic acid and rinsed in sterile deionized water as described previously (Paterson and Sim, 1999
). The seeds were then transferred aseptically onto discs of Tygan mesh at a density of approximately 45 seeds per disc. Individual discs were then placed over 1.0 l of deionized water within sterile culture vessels comprising of a round 2.0 l flat bottomed, wide-necked Duran flask (Schott Glaswerke, Mainz, Germany) covered by an upturned 95 mm diameter crystallizing dish. The discs of Tygan mesh were held at the level of the water surface by a polytetrafluoroethylene (PTFE) framework. The culture vessels were then placed within a controlled environment room (Conviron, Winnipeg, Canada) at 20 °C in the dark.
After 5–7 d in the dark the seeds had germinated, the water in the culture vessels was replaced, under aseptic conditions in a laminar airflow cabinet, with a nutrient solution sterilized by passing through a 0.2 µm cellulose nitrate filter (Whatman, Maidstone, UK). The composition of the complete nutrient solution was as described earlier (Thornton and Bausenwein, 2000) except that nitrogen was supplied either as 1 mol m-3 NH4NO3 (experiments 1 and 2) or 1 mol m-3 (NH4)2SO4 (experiment 3) and the final pH of all solutions were adjusted to 6.0 unless otherwise stated (experiment 3). The nutrient solutions were continuously aerated at a rate of 60 cm3 min-1 vessel-1 by passing air through an autoclaved PolyVENTTM 4 filter, 0.2 µm pore size (Whatman, Maidstone, UK) and sterile 1 mm internal diameter PVC tubing which was passed from outside the vessel, under the crystallizing dish ‘lid’ and into the nutrient solution. To ensure the airline remained in the solution, the end of the tube was weighted with a 1 cm length sleeve of additional PVC tubing.
Once germinated, plants were grown with a 16 h photoperiod of 550 µmol m-2 s-1 PAR at plant height. The temperature of the controlled environment room was adjusted to 12 °C during the light and 20 °C during the dark. The lower room temperature during the light period was to counteract radiant heat from the lights; the plants within their culture vessels experienced a constant temperature of 20 °C. Nutrient solutions were renewed aseptically every 7 d during the growth period. For all experiments, vessel positions were fully randomized within the controlled environment room. Prior to and during 15N labelling (see below) plants were subject to continuous light. As the start of the 15N labelling period in individual vessels was staggered, continuous light avoided interactions between the time of starting the labelling period and subsequent uptake of 15N label.
Experiment 1: Chemical speciation of N derived from Gly uptake
Fifteen vessels of plants were grown with the 16 h photoperiod as described above for 12 d. Light was then supplied continuously, and the plants allowed to grow for a further 2 d. Five replicate vessels were destructively harvested. In the remaining 10 vessels, the nutrient solution was replaced aseptically with one identical to that used for growth except all N was supplied as 2 mol m-3 Gly enriched with 15N to 49.1 atom%. Five replicate vessels were then harvested 3 h and 24 h after supplying the 15N-enriched Gly solution. At harvest, the plant roots were dipped in 1 mol m-3 CaSO4 at 4 °C for 1 min to remove any Gly adsorbed to the cell walls. Plants in each vessel were counted and separated into root, shoot and original seed material. The seeds were discarded; other material was weighed fresh. A subsample of root material was also weighed after being dried at 65 °C for 3 d. All shoot and the remaining root material was immediately frozen and stored at -80 °C. Amino acids were extracted from the plant material using a method adapted from Calanni et al. (Calanni et al., 1999). Frozen material was ground to a powder in liquid N2 using a mortar and pestle. The resultant powder was extracted with 80% (v/v) ethanol, 2 cm3 per 100 mg fresh weight, for 15 min at room temperature with occasional shaking. The solution was then centrifuged at 3500 g for 10 min. The supernatant was retained and the pellet resuspended in 80% ethanol (1 cm3 per 100 mg fresh weight) for 15 min then centrifuged once more at 3500 g for 10 min. The supernatants were combined, rotary evaporated to dryness at 40 °C, then resuspended in 10 cm3 deionized water. The water extract was purified using ion exchange SephadexTM (SP SephadexTM C25, Sigma) as described previously (Redwell, 1980). The 0.2 kmol m-3 NH4OH eluate from the SP SephadexTM C25 column containing the amino acids was rotary evaporated to dryness at 40 °C then resuspended in 1 cm3 deionized water. The concentration and 15N enrichment of the individual amino acids in the final water extract were determined by GC-MS as described earlier (Millard et al., 1998). Briefly, following addition of an internal standard of nor-valine, amino acids were converted to their t-butyldimethylsilyl derivatives, analysis of the derivatives was carried out by GC-MS in single ion recording mode (Trace 2000 series gas chromatograph linked to a Finnigan Trace mass spectrometer; ThermoQuest Limited, Manchester, UK). The derivatization used did not allow determination of Arg in the samples. The Arg content of six of the water extracts (one each, randomly chosen, of root and shoot extracts at 0 h, 3 h and 24 h) was checked using ion exchange chromatography with post column derivatization using ninhydrin. In all cases Arg contributed 2% or less of the amino acid N content of the sample.
The concentration and 15N enrichment of the amino acids determined by GC-MS was used to determine the labelled N content of each amino acid, derived from the uptake of Gly 15N, using equations described previously (Millard and Nielsen, 1989).
Experiment 2: Effects of defoliation and sugars on uptake of Gly-N
Twenty-five vessels of plants were grown with the 16 h photoperiod as described above for 12 d. Light was then supplied continuously, and the plants allowed to grow for a further 2 d. In all vessels, the nutrient solution was replaced aseptically with one identical to that used for growth except all N was supplied as 2 mol m-3 Gly enriched with 15N to 4.99 atom%. Five replicate vessels were assigned to each of five treatments: (1) plants left intact, (2) plants defoliated to 1.5 cm height, (3) plants defoliated and 25 mol m-3 sucrose added to the uptake solution, (4) plants defoliated and 50 mol m-3 glucose added to the uptake solution, and (5) plants defoliated and 50 mol m-3 3-O-methyl-D-glucopyranose added to the uptake solution. 3-O-methyl-D-glucopyranose (also referred to as 3-O-methylglucose) is an analogue of glucose that is absorbed, but not metabolized, by plants (Jang and Sheen, 1994; Bingham et al., 1996).
After 24 h, plants were harvested and separated as in experiment 1. Root and shoot material was dried at 65 °C for 3 d, weighed and ball milled. The total N and 15N concentrations were determined using a TracerMAT continuous flow mass spectrometer (Finnigan MAT, Hemel Hempstead, UK). The 15N enrichment was used to calculate the uptake of Gly-N from the 15N-labelled nutrient solution using equations described earlier (Millard and Nielsen, 1989).
Experiment 3: Uptake of Gly-N and ammonium N at varying pH
Forty-nine vessels of plants were grown with the 16 h photoperiod as described above for 16 d. Light was then supplied continuously, and the plants allowed to grow for a further 5 d. In 28 vessels, the nutrient solution was replaced aseptically with one identical to that used for growth except that all N was supplied as 2 mol m-3 Gly enriched with 15N to 5.02 atom%. In four replicate vessels each, the pH of the uptake solution was initially adjusted to 2.05, 3.46, 4.70, 5.94, 7.02, 7.79 or 9.43. Solution pH was adjusted by adding drops of either HCl or KOH as appropriate, initially at a concentration of 1 M then at a concentration of 0.1 M. In a further 21 vessels, the nutrient solution was replaced aseptically with one identical to that used for growth except all N was supplied as 1 mol m-3 (NH4)2SO4 enriched with 15N to 5.07 atom%. In three replicate vessels each, the pH of the uptake solution was initially adjusted as described above to 2.22, 3.47, 4.82, 5.99, 7.08, 8.02 or 9.52. After an uptake period of 24 h, plants were harvested and analysed as in experiment 2. The pH of all uptake solutions was measured at the end of the uptake period.
Statistics
Differences between treatments were assessed by analysis of variance using Genstat 5 Release 4.1©Lawes Agricultural Trust (IACR-Rothamsted). Results of allocation of uptake (percentage of uptake appearing in shoot) were subject to angular arc-sine transformation prior to analysis, as transformation did not alter the interpretation of results, untransformed data are presented for clarity.
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Chemical speciation of N derived from Gly uptake
In the free amino acid pool of root tissue, the majority of the labelled N derived from the uptake of Gly was present as Gly-N, Ser-N and to a lesser extent glutamine (Gln) N (Fig. 1
). Gly and Ser alone contributed 80±6% (mean±SD) of the total labelled N content of root free amino acids after a 3 h uptake period, this increased to 90±4% following 24 h of Gly uptake (Fig. 1). In roots, irrespective of the uptake period, more labelled N was present in the form of Ser alone than the sum of that present in all amino acids other than Gly and Ser (For 3 h P<0.05 and for 24 h P<0.01).
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In the shoot amino acid pool most labelled N, after a 24 h uptake period, was present in the form of Ser (Fig. 1
). However, the labelled N derived from Gly uptake was incorporated into a greater number of amino acids in the shoot compared with roots. In addition to the incorporation into Gly, Ser and Gln observed in roots, significant incorporation into asparagine (Asn), glutamic acid (Glu), aspartic acid (Asp), 
-aminobutyric acid (Gaba), and alanine (Ala) also occurred in shoots (Fig. 1).
Effects of defoliation and sugars on uptake of Gly-N
When sugars were absent from the uptake solution, defoliation reduced the uptake of Gly-N (P<0.01; Fig. 2A). The addition of either 25 mol m-3 sucrose or 50 mol m-3 glucose to the uptake solution of defoliated plants increased uptake of Gly-N compared with both defoliated plants without sugars and with undefoliated plants (P<0.01 in each case, Fig. 2A). The addition of 50 mol m-3 3-O-methyl-D-glucopyranose did not cause any change in the uptake of Gly-N by defoliated plants (P>0.05; Fig. 2A). When uptake of Gly-N was altered through manipulation of defoliation and sugar supply, plants with the greatest uptake (i.e. defoliated plants supplied with either sucrose or glucose) had the lowest allocation of uptake to shoot material (P<0.05, Fig. 2B).
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), undefoliated; (
), defoliated; (
), defoliated and 25 mol m-3 sucrose added to the uptake solution; (•), defoliated and 50 mol m-3 glucose added to the uptake solution; or (
), defoliated and 50 mol m-3 3-O-methyl-D-glucopyranose added to the uptake solution.
Uptake of Gly-N and ammonium N at varying pH
Although the uptake solutions used to investigate the effects of pH on uptake of Gly and contained phosphate buffer (Thornton and Bausenwein, 2000) changes in pH did occur during the uptake period. The maximum changes observed were a reduction of 0.6 pH units for Gly uptake and a reduction of 2.0 pH units for uptake. Most observed changes were considerably less than these maxima. All reported pH values refer to median values between the initial and final pH of the uptake solutions.
An increased pH of the uptake solution from 2.1 to 3.5 caused a 2.9-fold increase in the uptake of Gly-N (P<0.01; Fig. 3A). Further increases in pH above 3.5 resulted in decreased Gly-N uptake, such that uptake at pH 3.5 was greater than at pH 5.7, 6.9 and pH 7.5, which in turn was greater than at pH 9.2 (P<0.05 in all cases; Fig. 3A). In contrast to manipulation of Gly-N uptake through the addition of sugars; when Gly-N uptake was altered through differences in pH of the uptake solution, plants with the greater uptake of Gly-N allocated a greater percentage of their uptake to shoot material (P<0.001; Fig. 3B).
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Overall, an increased pH of the uptake solution from 2.2 to 9.1 caused an increase in
uptake by plants of L. perenne (P<0.001; Fig. 4A). However, the increases in uptake occurred mainly at either end of the pH range studied. Changing pH over the range 3.4–7.5 caused no alteration in uptake (P>0.05; Fig. 4A). At pH 6, uptake of N in the form of was approximately 5 times that of N uptake in the form of Gly, at pH 9 the relative uptake of -N had increased to over 13 times that of Gly-N (Figs 3A, 4A). Allocation of -N to shoot material increased (P<0.001) with increased uptake (Fig. 4B).
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The presence of 15N label in the amino acid pool of roots and shoot following incubation of the roots with 15N-enriched Gly proves that non-mycorrhizal plants of L. perenne are capable of acquiring N from Gly. In plants Ser can be synthesized directly from Gly and the two amino acids are readily inter-converted (Keys, 1980
; Bourguignon et al., 1999). An assumption was made that labelled Ser-N observed in the root amino acid pool was previously metabolized from labelled Gly-N. The large proportion of N derived from Gly uptake in the root amino acid pool which either remained as Gly-N or was in the form of Ser suggests a minimum of 80% of Gly-N uptake was as an intact Gly molecule. In the sterile solution culture used, microbial degradation of Gly to outside the root would not have occurred. However, the possibility still exists that Gly could have been degraded outside the plant if the appropriate enzymes were exuded from roots. The uptake of any 15 formed in this manner would have resulted in the 15N label initially appearing in the root amino acid pool as Gln through the action of glutamine synthetase (GS). In Zea mays 15N-labelled Gln was formed within minutes of supplying plants 15 (Lee and Lewis, 1994). If the formation of 15 in the sterile nutrient solution occurred and its subsequent uptake was significant and GS activity was low, 15 may have accumulated in the L. perenne roots. However, GS activity reported for non-mycorrihizal roots of L. perenne (61 µmol h-1 g-1 FW, Faure et al., 1998) would appear sufficient to avoid a large build up of given the rates of Gly uptake currently reported.
The apparent deviation from 100% uptake of Gly-N as intact Gly is more likely to have occurred by post-uptake processes. Any metabolism of Gly and Ser to Gln in the roots would cause an underestimate in the percentage of Gly-N uptake that occurred as intact Gly. Conversely, preferential losses of amino acids other than Gly and Ser from the root amino acid pool either by incorporation into protein or transport to the shoot would cause an overestimate of this value. However, the large amount of labelled Ser found in the shoot of L. perenne indicates that this amino acid was readily transported. Indeed Ser, which had been derived from uptake of Gly, was suggested to be preferentially transported out of the roots of Hakea species rather than undergoing further metabolism (Schmidt and Stewart, 1999). Root–shoot–root cycling of amino acids in the manner described previously (Cooper and Clarkson, 1989) could also have potentially affected the estimate of intact Gly uptake in either direction. The extent to which these post-uptake processes affected the estimate of the proportion of Gly-N uptake which occurred as intact Gly during a 3 h period of uptake is unknown.
Due to the different techniques used direct comparison is difficult, however it is not altogether surprising that the current estimate of 80% of Gly-N uptake by L. perenne being as intact Gly, is greater than previous estimates in soil with competing microbes, of 19% and 64% by P. pratense and D. flexuosa, respectively (Näsholm et al., 1998, 2000). In soil systems large uncertainties exist in relation to the degree of Gly transformation prior to uptake, comparison of intact Gly uptake using 15N labelling coupled with GC-MS in both sterile and soil based systems may allow the degree of such transformations to be assessed.
In planta, Ser can be synthesized from Gly by two reactions (Bourguignon et al., 1999). The first combines the action of serine hydroxymethyltransferase (SHMT) with the glycine decarboxylase complex (GDC); essentially two moles of Gly produce one mole of Ser and one mole of ammonia (see Bourguignon et al., 1999, for further details). The ammonia being rapidly refixed by glutamine synthetase and further metabolized into other amino acids. Hence if this pathway had been used it would be expected that an equal number of moles of Ser and ‘other’ amino acids would have been 15N-labelled. The second utilizes serine:glyoxylate aminotransferase (SGAT) by which one mole of Gly combines with hydroxypyruvate to produce one mole of Ser and glyoxylate (Bourguignon et al., 1999). The greater content of labelled N as Ser than that in all amino acids other than Gly and Ser in the root amino acid pool following a 3 h uptake of Gly, suggests SGAT must have been used to synthesize Ser to some degree. The potential problems in estimating intact Gly uptake highlighted above, such as possible 15 accumulation in roots if GS activity is low must again be considered pertinent. Through the use of inhibitors it has been concluded (Schmidt and Stewart, 1999) that in roots of Hakea species SHMT was inactive and the catabolism of Ser from Gly was via SGAT.
The uptake of many amino acids by plants exhibit distinct pH optima which, in the main, fall between pH 4 and pH 6 (Soldal and Nissen, 1978; van Bel et al., 1981). The uptake of Gly by leaves of Hordeum vulgare (barley) did exhibit an optimum pH of 5.8 (Lien and Rognes, 1977), in contrast, Gly uptake by leaf protoplasts of Pisum sativum (pea) showed little change over the pH range 2–9 (Dureja et al., 1984). The pattern of change in the permeability coefficient of Gly efflux across unilamellar vesicles with pH (Chakrabarti and Deamer, 1992) matches closely the effect of pH on whole plant Gly uptake in the present study. The current result of decreasing Gly uptake with increasing pH over the range 3.5–9.2 is consistent with Gly uptake, in common with other amino acids, being via a proton symport (Bush and Langston-Unkefer, 1988; Li and Bush, 1991). It is interesting to note that the maximum rate of Gly uptake observed at pH 3.5 did not coincide with the isoelectric point of Gly (pH 6.0). The reduction of Gly uptake below pH 3.5 may have been due to membrane damage as uptake was also substantially reduced at the lowest pH.
Amino acid proton symports are considered dependent upon an actively generated proton electrochemical potential difference. Evidence that this is the case for uptake of Gly by roots comes from inhibition of Gly uptake by the sedge Kobresia myosuroides, on reduction of the trans-membrane electrochemical potential either by bubbling N2 through the nutrient solution or with the addition of the protonophore carbonyl cyanide m-chlorophenylhydrazone (Raab et al., 1996). The currently observed response of Gly uptake to defoliation and to externally applied sugars or a sugar analogue incapable of metabolism indicates Gly uptake by roots of L. perenne is energy dependent. These results also indicate, as previously suggested for NO3-uptake by L. perenne (Clement et al., 1978), that the uptake of Gly is dependent on current photosynthate. The allocation of N between root and shoot, derived from the uptake of Gly is clearly flexible, current results indicate that in part this is determined by the root carbohydrate status.
The considerably lower uptake rate of Gly compared with at equimolar concentrations of N, coupled with a greater availability of than Gly in soil following urine deposition (Thomas et al., 1988), suggest Gly uptake is unlikely to make a large contribution to the total N acquisition of L. perenne plants at this time. The large increase in soil pH following urine deposition (Thomas et al., 1988) would serve to emphasize the greater uptake of relative to Gly, especially if pH were to be increased above pH 7.5. A similar shift in relative uptake of Gly and with changing pH was inferred by Schiller et al. who concluded most uptake of Gly by the aquatic plant Chamaegigas intrepidus, from pools containing both Gly and , would occur when pH was lowest (Schiller et al., 1998). In the field L. perenne will be mycorrhizal, whilst association with AM fungi increases uptake of amino acids (Cliquet et al., 1997), it also increases uptake of (Johansen et al., 1993). Whether the relative acquisition of Gly and by L. perenne changes following AM fungal infection is unknown.
In the sedges Eriophorum vaginatum, Carex aquatilis (Schimel and Chapin, 1996) and K. myosuroides (Raab et al., 1996) and in the grass P. pratense (Näsholm et al., 2000) the relative uptake of Gly compared with inorganic N uptake was greater than that observed in this study. These differences are unlikely to be due to differences in mycorrhizal status, as both the K. myosuroides (used by Raab et al., 1996) and the L. perenne in the current study were non-mycorrhizal. These differences may reflect actual species differences in relative uptake of the different forms of N (Schmidt and Stewart, 1999; Falkengren-Grerup et al., 2000). A second possibility is that in soil studies, but not in solution culture, will be preferentially adsorbed onto soil surfaces, reducing its availability to roots. Alternatively, the differences in relative uptake of Gly and inorganic N may reflect the degree to which plants had prior exposure to Gly. The E. vaginatum, C. aquatilis and P. pratense (Schimel and Chapin, 1996; Näsholm et al., 2000) were grown in soil, whilst the plants of K. myosuroides (Raab et al., 1996) were supplied Gly during growth. The plants of L. perenne in the current study had no exposure to Gly prior to the uptake period. Induction of Gly uptake may occur in a manner analogous to that observed for inorganic N (Ourry et al., 1997), though induction of proline uptake by Ricinus communis was not evident following a 19 h preincubation period with proline (Schobert and Komor, 1987).
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Acknowledgments |
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The Scottish Executive Rural Affairs Department funded this work. I thank Mr A Hepburn and Mrs MM Procee of the Macaulay Land Use Research Institute (MLURI) for GC-MS analysis, Mr DS Brown of the Rowett Research Institute for ion exchange chromatography analysis, Mrs MR Tyler of MLURI for skilled technical assistance and Dr E Paterson for helpful comments.
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Notes |
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1 Fax: +44 1224 311556. E-mail: b.thornton{at}mluri.sari.ac.uk
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