Supplemental Figure 1

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Supplemental Figure 1
Supplemental Figure 1. Levels of nitrate,
carbohydrates and metabolites involved in nitrate
assimilation, and the medium pH. Starch and the
sum of Glucose, Fructose and Sucrose are
expressed as µmole hexose equivalents per gram
fresh weight (µmoles HEq gFW-1). Seedlings were
grown in liquid culture with full nutrients
including 0.5 sucrose in continuous light for 7
days, and then for a further 2 days on full
nutrient medium, or on low N medium quickly
leading to N starvation (see Materials and
Methods). Samples were taken two days later from
seedlings on full medium and seedlings on
N-deficient medium, and from seedlings that had
been incubated on N-deficient medium for 2 days
before adding 3 mM KNO3 for 30 min, 3 h and (see
Figures 4 and 5) 8 and 24 h. Control N-deficient
seedlings were provided with 3mM KCl for 30 min,
3, 8 and 24 h. Metabolite levels in N-starved
seedlings prior to NO3- addition are shown as
cross-hatched bars, those of KNO3-fed ones as
gray bars, the KCl controls as white bars, and
those of N-replete seedlings as black bars.
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Supplemental Figure 2
Supplemental Figure 2. Comparison of gene
expression signals and signal ratios, as
determined by Affymetrix technology.   (A)
Double-logarithmic plot of gene expression
signals of two biological replicates of N-starved
seedlings 30min after NO3- addition. Linear
regression, including signals called present as
well as absent, yields a correlation
coefficient (R2) of 0.923. (B) Distribution plot
of signal ratios between the two biological
replicates. (C) Double-logarithmic plot of gene
expression signals of N-starved seedlings prior
and after 3hrs 3mM KCl addition. Linear
regression, including signals called present as
well as absent, yields a correlation
coefficient (R2) of 0.924. (D) Distribution plot
of signal ratios between the two samples shown in
(C). (E) Double-logarithmic plot of gene
expression signals of N-starved seedlings at time
zero and N-starved seedlings 3 h later without
any addition. Linear regression, including
signals called present as well as absent,
yields a correlation coefficient (R2) of
0.917. (F) Distribution plot of signal ratios
between the two samples shown in (E).
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Supplemental Figure 2. continued
Supplemental Figure 2. Continued.   (G, H)
Comparison of transcript ratios after 3 h 3mM
KNO3 or 3 h 3mM KCl treatment of N-starved
seedlings, as determined by ATH1 analysis. The
ratios are shown for 335 genes that showed
gt3-fold increase or decrease after 3 h nitrate
addition (black bars). The expression ratios
determined after 3 h KCl addition are shown as
grey bars. (I) Double logarithmic plot of the
same data.
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Supplemental Figure 3
Supplemental Figure 3. BIN description for the
metabolic overview plot (see Figure 2).
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Supplemental Figure 4A
Supplemental Figure 4A. Expression of NRT2
family genes and a gene encoding a Chlamydomonas
NAR2 homolog. Bars (from left to right) in each
group of five per gene represent ATH1 expression
levels standard error (n2-3) in full nutrition
(N), in N-starvation (-N), after 30min 3mM KNO3
addition, after 3h 3mM KNO3 addition, and 3mM KCl
addition, respectively. Asterisks denote
expression levels that were called absent by
Affymetrix software.
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Supplemental Figure 4B
Supplemental Figure 4B. Expression of genes
involved in transport in seedlings 30min after
addition of nitrate. The results are shown as
changes relative to the level in N-deficient
seedlings.
Metals
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Supplemental Figure 4C
Supplemental Figure 4C. Expression of genes
involved in transport in seedlings 3 hrs after
addition of nitrate. The results are shown as
changes relative to the level in N-deficient
seedlings.
Metals
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Supplemental Figure 4D
Supplemental Figure 4D. Expression of genes
involved in transport, in N-replete seedlings.
The results are shown as changes relative to the
level in N-deficient seedlings.
Metals
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Supplemental Figure 5A
Supplemental Figure 5A. Expression of genes
genes involved in glycolysis, the tricarboxylic
acid cycle and mitochondrial electron transport,
resolved to the single enzyme level , in
seedlings 30min after addition of nitrate. The
results are shown as changes relative to the
level in N-deficient seedlings.
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Supplemental Figure 5B
Supplemental Figure 5B. Expression of genes
genes involved in glycolysis, the tricarboxylic
acid cycle and mitochondrial electron transport,
resolved to the single enzyme level , in
seedlings 3hrs after addition of nitrate. The
results are shown as changes relative to the
level in N-deficient seedlings.
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Supplemental Figure 5C
Supplemental Figure 5C. Expression of genes
genes involved in glycolysis, the tricarboxylic
acid cycle and mitochondrial electron transport,
resolved to the single enzyme level , in
N-replete seedlings. The results are shown as
changes relative to the level in N-deficient
seedlings.
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Supplemental Figure 6
Supplemental Figure 6. Expression of genes
involved in RNA and protein synthesis, in
seedlings 30min after addition of nitrate . The
results are shown as changes relative to the
level in N-deficient seedlings.
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Supplemental Figure 7A
Supplemental Figure 7A. Expression of members of
large gene families, in seedlings 30min after
addition of nitrate. The results are shown as
changes relative to the level in N-deficient
seedlings.
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Supplemental Figure 7B
Supplemental Figure 7B. Expression of members of
large gene families, in seedlings 3hrs after
addition of nitrate. The results are shown as
changes relative to the level in N-deficient
seedlings.
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Supplemental Figure 7C
Supplemental Figure 7C. Expression of members of
large gene families, in N-replete seedlings. The
results are shown as changes relative to the
level in N-deficient seedlings.
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Supplemental Figure 8A
Supplemental Figure 8A. Expression of genes
involved in cellular regulation, in seedlings
30min after addition of nitrate. The results are
shown as changes relative to the level in
N-deficient seedlings.
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Supplemental Figure 8B
Supplemental Figure 8B. Expression of involved in
cellular regulation, in seedlings 3hrs after
addition of nitrate. The results are shown as
changes relative to the level in N-deficient
seedlings.
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Supplemental Figure 8C
Supplemental Figure 8C. Expression of involved in
cellular regulation, in N-replete seedlings. The
results are shown as changes relative to the
level in N-deficient seedlings.
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Supplemental Figure 9
Supplemental Figure 9. Comparison of gene
expression ratios after short-term nitrate
re-addition as determined by Affymetrix
technology, using data from the present paper and
data from (Wang et al., 2003).   (A) Expression
ratios from both studies for all 22750 probe-sets
plotted on a double logarithmic scale. (B)
Distribution plot of the double ratios (ratio X /
ratio Y) for roots (black bars) and shoots (grey
bars). Of the 22750 probe sets on the ATH1 array
more than 98.5 gave ratios that were less than
2-fold different, and only nine probe sets
(lt0.04) gave ratios that were 5-fold or more
different when our materials were compared to the
roots analyzed in Wang et al. (2003). (C)
Gene-by-gene comparison of transcript ratios for
176 genes that showed a gt3-fold increase. Cyan
and blue bars represent ratios calculated for
roots and shoots, respectively, from the data of
Wang et al., (2003). Red bars represent ratios
calculated from data of whole seedlings published
in the present paper.
A
B
C
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Supplemental Figure 10A.
Supplemental Figure 10A. MAPMAN metabolic
overview plot created with Shoot data from Wang
et al. (2003).
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Supplemental Figure 10B
Supplemental Figure 10B. MAPMAN metabolic
overview plot created with Root data from Wang et
al. (2003).
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Supplemental Figure 11A
Supplemental Figure 11A. MAPMAN transport
overview plot created with Shoot data from Wang
et al. (2003).
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Supplemental Figure 11B
Supplemental Figure 11B. MAPMAN transport
overview plot created with Root data from Wang et
al. (2003).
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Supplemental Figure 12A
Supplemental Figure 12A. MAPMAN gene family
overview plot, created with Shoot data from Wang
et al. (2003).
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Supplemental Figure 12B
Supplemental Figure 12B. MAPMAN gene family
overview plot, created with Root data from Wang
et al. (2003).