Transcript NPH_3580_sm_FigS1-S6

(a)
pGlcT
TPT
rRNA
rRNA
(b)
pGlcT
rRNA
(c)
TPT
pGlcT
rRNA
rRNA
TGG
Wild-type
TGA
Homozygous
mutant
Fig. S1 Characterization of homozygous plastidic sugar transporter mutants. The null
expression of the corresponding genes in the single (a) and double (b) mutants was confirmed
by Northern blot analysis. Ethidium bromide-stained rRNAs are shown as the respective
controls. (c) The homozygous mex1 allele in the pglct-1/mex1 double mutant was confirmed
by sequencing, because mex1 is a single point mutation.
8000
pGlcT
6000
4000
2000
Signal intensity
0
3000
Leaf
Root
Stem
MEX1
Flower
Leaf
Root
Stem
TPT
Flower
Leaf
Root
Stem
Flower
2000
1000
0
40000
30000
20000
10000
0
Fig. S2 Digital Northern analysis of plastidic sugar transporters. Microarray data for adult
rosette leaf, roots, inflorescence stem and flower of Arabidopsis were obtained from GEO
(Gene Expression Omnibus) database at the Genvestigator site (https://www.genvestigator.com)
and used for the digital Northern analysis.
Maltose content (μmol/gFW)
10
8
End of day
End of night
*
* *
6
4
*
2
0
Fig. S3 Maltose content of mutant and wild-type plants at the end of the day and night.
Maltose content was determined in the mature leaves of 4-week-old single (pglct-1, pglct-2
and mex1) and double (pglct-1/mex1) mutants at the end of the day and night. Each point
represents the mean ( SD) from four different measurements within each line. Asterisks
indicate significant differences (p < 0.05, t-test) between mutant and wild-type plants. FW;
fresh weight.
No sugar
+ Glucose
+ Maltose
WT
pglct-1/mex1
tpt-2/mex1
Fig. S4 Recovery of the defective phenotypes of the pglct-1/mex1 and tpt-2/mex1 mutants by
supplying external sugars, glucose and maltose. Mutant and wild-type plants were grown on
agar plates containing Gamborg B5 media complemented with 2% of each sugar. The pictures
shown are of 25-day-old mutant and wild-type plants.
(a)
(b)
4
WT
pglct-1
pglct-2
tpt-2
mex1
*
3
*
2
*
*
1
5
Glucose content (mol/gFW)
Glucose content (mol/gFW)
5
**
WT
pglct-1/tpt-2
pglct-1/mex1
tpt-2/mex1
4
*
3
2
*
1
*
0
0
0
4
8
12
16
20
0
24
4
(c)
(d)
2.0
Fructose content (mol/gFW)
3.0
WT
pglct-1
pglct-2
tpt-2
mex1
2.5
*
1.5
*
1.0
8
12
16
20
24
Time of the day (h)
Time of the day (h)
Fructose content (mol/gFW)
*
*
0.5
3.0
WT
pglct-1/tpt-2
pglct-1/mex1
tpt-2/mex1
2.5
*
2.0
*
1.5
1.0
*
*
*
0.5
*
0.0
0.0
0
4
8
12
16
Time of the day (h)
20
24
0
4
8
12
16
20
24
Time of the day (h)
Fig. S5 Diurnal changes in glucose and fructose levels in the plastidic transporter mutants.
Glucose (a and b) and fructose (c and d) contents were determined in the mature leaves of 4week-old single (a and c) and double (b and d) mutants during the diurnal cycle. Each point
represents the mean ( SD) from five different measurements within each line. Asterisks
indicate significant differences (p < 0.1, t-test) between mutant and wild-type plants. The
black shading in the upper bar indicates the dark period. FW; fresh weight.
ISA3
BAM3
DPE1
DPE2
SEX1
PHS1
TUB
Fig. S6 Effects of plastidic sugar transporter defects on the expression of starch degradationrelated enzymes. Expression levels of starch degradation-related genes, isoamylase 3 (ISA3),
β-amylase 3 (BAM3), disproportionating enzyme 1 (DPE1), DPE2, glucan-water dikinase
(SEX1), and glucan phosphorylase (PHS1) in the middle of day were analyzed by RT-PCR.
Total RNA was isolated from the leaves collected at the middle of day and night of
Arabidopsis plants grown for 4 weeks. The tubulin 2 gene (TUB) was amplified as a control.