1. Supplemental Fig. S1 A B
AtMYBS1 3 57
128
180
314 aa
AtMYBS2 7 58
112
164
298 aa
Fig. S1. Sequence alignment and domain structure of the MYBS1 and MYBS2 proteins.
(A) Amino acid sequences for MYBS1, MYBS2, and OsMYBS1were compared using the GeneDoc program. Identical residues are shown in black and similar residues in gray. The most conserved MYB DNA-binding domain and weakly conserved MYB-like DNA-binding domain are highlighted by a black line and a gray line, respectively. The high conserved SHAQKYF motif is indicated by the arrow. (B) The structures of the MYBS1 and MYBS2 proteins. Both MYBS1 and MYBS2 contain a conserved MYB DNA-binding domain and a weakly conserved MYB-like DNA-binding domain, which is located at the N-terminus of the protein.

2. Supplemental Fig. S2 A B C D
LBa1
S2LP
ATG
TAA
mybs2
S2RP
LB3
S1RP
S1LP
TGA
mybs1
S1F
S1R
S2F
S2R A
MYBS1
MYBS2
ACT1
mybs1
mybs2 WT B C D WT
mybs1
mybs2 WT
mybs1
mybs2
Fig. S2. Identification of mybs1 and mybs2 mutant lines. A, Schematic diagram represents structure of MYBS1 and MYBS2 genes. Block boxes and gray boxes represent coding sequences and untranslated sequences, respectively. The T-DNA insertion positions are indicated by open triangles. The positions of the primer sequences used for PCR genotyping the mybs1 mutant alleles and the mybs2 are marked with arrows. The primers S1RP and S1LP were used to screen the mybs1 mutants. The primers S2RP and S2LP were designed specifically for screening of mybs2 mutant. The primers S1F combined with S1R were used to detect the full-length of MYBS1 mRNA. The primer set S2F and S2R was used to detect the MYBS2 full-length mRNA. B, RT-PCR analysis for MYBS1 and MYBS2 mRNA in the wild type (WT), mybs1and mybs2 mutants. RNAs were isolated from 2-weeks-old seedlings and subjected to RT-PCR analysis. Arabidopsis Act1 was used as an internal control. C and D, Phenotypes of WT, mybs1, and mybs2 growth in the pots under normal conditions for 28 days (C) and 45 days (D).

3. Supplemental Fig. S3 Col-0 Col-3 mybs2 mybs1 4% Glucose for 6 days
Fig. S3. Phenotypes of WT, mybs1, and mybs2 growth in 4% (left) and 5% (right) glucose containing medium for 6 days (left) and 14 days (right).

4. Supplemental Fig. S4
BAMY1
BAMY3
AMY3
SUS1
SUS2
SUS3 WT
mybs1
mybs2
Relative level M G A B * * * * * * * * * * * * * * * * *
Fig. S4. Expression of carbohydrate catabolic related genes in seedlings of WT, mybs1, and mybs2. A, Total RNAs isolated from seven-day-old wild-type seedlings grown on 4% glucose- or mannitol-containing medium were subjected to qRT-PCR analyses using primers specific for AMY3, BAMY1, BAMY3, SUS1, SUS2, and SUS3. The UBQ10 gene was used as an internal control. Gene expression levels were relative to that in seedlings grown on 4% mannitol containing medium, which was set as 1. An asterisk indicates a significant difference from the seedlings grown on 4% mannitol containing medium (Student’s t test: p <0.05). B, Total RNAs isolated from seven-day-old wilid-type seedlings grown on 4% glucose-containing medium were subjected to qRT-PCR analyses using primers specific for AMY3, BAMY1, BAMY3, SUS1, SUS2, and SUS3. The UBQ10 gene was used as an internal control. Gene expression was relative to that in wild-type plants, which was set as 1. An asterisk indicates a significant difference from the wild-type plants (Student’s t test: p <0.05).

5. Supplemental Fig. S5
Fig. S5. Expression of ABA biosynthesis and ABA signaling genes in WT, mybs1, and mybs2 seedlings. Total RNAs isolated from seven-day-old seedlings grown on 4% glucose-containing medium were subjected to semi-quantitative RT-PCR analyses using primers specific for ABA1, NCED9, AAO3, ABI3, ABI4, and ABA5. The UBQ10 gene was used as an internal control.

6. Supplemental Fig. S6
Fig. S6. Putative cis-acting elements present in promoters of sugar- and ABA-related genes that change expression in mybs1 and mybs2. Positions of the TA box and TA-like boxs within 1 kb upstream of the putative transcription initiation site indicated by color-coded boxes, and their nucleotide sequences are shown at the bottom of the graph. Horizontal arrows indicate the orientation of the TA box and TA-like boxs in the promoter region. +1 indicate the position the putative transcription initiation site.