Plant Science 160 2001 219 – 228
Differential effect of amino acids on nitrate uptake and reduction systems in barley roots
Muhammad Aslam, Robert L. Travis , D. William Rains
Department of Agronomy and Range Science, Uni6ersity of California, One Shields A6enue, Da6is, CA
95616
-
8515
, USA Received 29 June 2000; received in revised form 6 August 2000; accepted 6 August 2000
Abstract
This study was conducted to determine whether the inhibition of nitrate reductase activity NRA; EC 1.6.6.1 in barley Hordeum 6ulgare L. var. CM-72 roots by the amino acids glutamic, aspartic, glutamine and asparagine is a direct effect or
indirect due to inhibition of the NO
3 −
uptake system. Roots of 8-day-old intact seedlings were supplied with the amino acids I mM individually either with NO
3 −
0.1 or 10 mM or roots were pretreated with the amino acids and then supplied with NO
3 −
only. Nitrate uptake was determined by following NO
3 −
depletion from the uptake solution containing 0.1 mM NO
3 −
. All the amino acids inhibited the increase in NO
3 −
uptake similarly 50 – 60 when the roots were supplied with 0.1 mM NO
3 −
. Pretreatment with glutamic and aspartic acids was more inhibitory 70 – 80 than with glutamine and asparagine 30. The
amino acids partially inhibited 35 the induction of NRA in roots supplied with 0.1 mM NO
3 −
; however, no inhibition occurred at 10 mM NO
3 −
. Likewise, pretreatment with glutamic or aspartic acid inhibited the induction of NRA at 0.1 mM NO
3 −
but not at 10 mM NO
3 −
. In contrast, pretreatment with glutamine or asparagine had no effect on the subsequent induction of NRA, even at 0.1 mM NO
3 −
. The results suggest that, at low NO
3 −
supply, the inhibition of induction of NRA by the amino acids is a result of the lack of substrate availability due to inhibition of the NO
3 −
uptake system. © 2001 Elsevier Science Ireland Ltd. All rights reserved.
Keywords
:
Amino acids; Barley; Nitrate uptake; Nitrate reductase; Roots www.elsevier.comlocateplantsci
1. Introduction
Nitrate is the most common form of inorganic N utilized by those higher plants, which lack
N
2
-fixing symbioses [1]. The assimilation of NO
3 −
into organic N requires the participation of several specific proteins [2]. First NO
3 −
, absorbed by the plasma membrane-bound transporter, is reduced
to NH
4 +
by nitrate reductase NR and nitrite reductase [3,4]. NH
4 +
is then incorporated into amino acids by the glutamine synthetase-glu-
tamine-2-oxoglutarate amidotransferase
GS- GOGAT enzyme system [5], giving rise to
glutamine Gln and ultimately other amino acids and their metabolites [6]. Free amino acids are the
precursors by which N is transferred between cells and organs [7]. Both NO
3 −
uptake and reduction processes are substrate inducible and are regulated
by endogenous metabolites including amino acids [3,4].
A rapid cycling of amino acids between roots and shoots occurs whether NO
3 −
assimilation is localized mainly in shoots [8 – 10] or roots [11].
Plants that exhibit low rates of NO
3 −
assimilation in roots export most of the absorbed NO
3 −
to shoots where it is reduced and incorporated into
amino acids. The amino acids from the shoots are
Abbre6iations
:
Asp, aspartic acid; Asn, asparagine; DON, dia- zoacetyl-
DL
-norleucine methyl ester; Glu, glutamic acid; Gln, glu- tamine; GOGAT, glutamine-2-oxoglutarate amidotransferase; GS,
glutamine synthetase; HPLC, high-performance liquid chromatogra- phy; IHATS, inducible high affinity transport system; LATS, low
affinity transport system; NRA, nitrate reductase activity; PPFD, photosynthetic photon flux density; SE, standard error.
Corresponding author. Tel.: + 1-530-7526162; fax: + 1-530- 7524361.
E-mail address
:
rltravisucdavis.edu R.L. Travis. 0168-945201 - see front matter © 2001 Elsevier Science Ireland Ltd. All rights reserved.
PII: S 0 1 6 8 - 9 4 5 2 0 0 0 0 3 9 1 - 5
then translocated to roots via the phloem [12 – 14]. Thus, plant roots may be exposed internally to
elevated levels of amino acids. The pool of amino acids cycling between the root and shoot is consid-
ered to serve as a signal for the plant internal N status [9,15,16]. The amino acids, whether accu-
mulated in the plant internally or supplied exter- nally, usually down regulate the induction of NO
3 −
uptake [2,15,17 – 20] and reduction systems [21 – 26].
Regulation of induction of the NO
3 −
uptake and reduction systems by reduced N compounds has
been attributed to feed-back inhibition [26,27]. Since the induction of both of these systems is
dependent upon the availability of NO
3 −
in the tissue [28], the N metabolites may affect the induc-
tion processes by affecting the availability of the substrate. Sivasankar et al. [26] observed that Gln
and asparagine Asn inhibited the induction of NR activity NRA in corn roots at both low and
high external NO
3 −
supply. They concluded that inhibition was not the result of altered NO
3 −
up- take, but was due to the direct effect of these
metabolites at the transcription level. In contrast, we recently observed that Gln partially inhibited
the induction of NRA in barley roots at lower, but not at higher, exogenous NO
3 −
concentrations M. Aslam, R.L. Travis, and D.W. Rains, unpublished
results. At low NO
3 −
concentrations uptake is mediated by the inducible high affinity transport
system IHATS; whereas at high NO
3 −
most ab- sorption is via the low affinity transport system
LATS [29]. While the induction of IHATS is inhibited by N metabolites [2,15,17,20,30], LATS
is a constitutive system [29,31] and may not be affected by amino acids. The induction of NRA in
both roots [32] and shoots [33] is regulated by NO
3 −
flux. Thus, the inhibition of induction of NRA by N metabolites in barley roots at lower
NO
3 −
supply may be due to decreased NO
3 −
up- take rather than a direct effect on the reduction
system. In the present study the role of glutamic acid
Glu, aspartic acid Asp, Gln and Asn in the induction of the NO
3 −
uptake system and of NRA in barley roots was investigated. These amino
acids were selected because in many plant species they are exported from the leaves to the roots via
the phloem in considerable amounts [34 – 37]. The results show that while these amino acids directly
inhibited enhancement of the activity of NO
3 −
uptake system, independent of NO
3 −
availability, inhibition of the induction of NRA is due to a
decrease in NO
3 −
uptake resulting in decreased NO
3 −
availability.
2. Materials and methods
