1. MYB34, MYB51, and MYB122 Distinctly Regulate Indolic Glucosinolate Biosynthesis in Arabidopsis thaliana 
Frerigmann Henning , Gigolashvili Tamara  
Molecular Plant 
Volume 7, Issue 5, Pages (May 2014)
DOI: /mp/ssu004
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

2. Figure 1 Biosynthesis of Indolic Glucosinolates.
The biosynthesis of tryptophan-derived indolic GSL and their relation to indole-3-acetic acid (IAA), camalexin, and indolecarboxylic acids is depicted. The known Arabidopsis thaliana genes involved in these steps are indicated. The figure is adapted from Sønderby et al. (2010b). IAOx, indole-3-ylacetaldoxime; I3M, indole-3-ylmethyl-GSL; 4HO-I3M, 4-hydroxy-indole-3-ylmethyl-GSL; 4MO-I3M, 4-methoxy-indole-3-ylmethyl-GSL; 1HO-I3M, 1-hydroxy-indole-3-ylmethyl-GSL; 1MO-I3M, 1-methoxy-indole-3-ylmethyl-GSL. Multiple enzymatic steps are indicated with interrupted arrows.
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

3. Figure 2
MYB34, MYB51, and MYB122 Are Indispensable for IG Biosynthesis under Standard Growth Conditions.
(A) The positions of T-DNA insertions in myb34 (WiscDsLox424F3), myb122–1/MYB122–1D (Salk_039228), and myb122–2 (WiscDsLoxHs206_04H) mutant lines are given. Boxes indicate exons, lines indicate introns.
(B) The indolic glucosinolate (IG) content of single and multiple myb mutants. For glucosinolate (GSL) analysis, leaves of 6-week-old plants were harvested and the three major IGs (I3M, 4MO-I3M, and 1MO-I3M) for wild-type (Col-0) and various myb knockouts are shown.
(C) Levels of I3M, 4MO-I3M, and 1MO-I3M relative to wild-type (Col-0 = 100%) and the sum of all GSL, IGs and aliphatic glucosinolates (AGs) is shown for myb34 myb51 myb122–2 and cyp79b2 cyp79b3 mutants. Results are means ± SE from three independent cultivations with four biological replicates (n = 12). Values marked with asterisks are significantly different from Col-0 (Student’s t-test; p < 0.05).
(D) Transcript levels of IG biosynthetic genes in 6-week-old leaves (Col-0 = 1). Data are means ± SE from three independent cultivations with three biological replicates (n = 9). Values marked with asterisks are significantly different from Col-0 (Student’s t-test; p < 0.05).
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

4. Figure 3
MYB34, MYB51, and MYB122 Play Different Roles in IG Biosynthesis in Shoots and Roots.
(A, C) Relative expression levels of MYB34, MYB51, and MYB122 in shoots (A) and roots (C) of wild-type (Col-0) plants. Expression data are presented relative to Actin2. Data are means ± SE from two independent cultivations with three biological replicates (n = 6).
(B, D) IG levels of single myb34, myb51, myb122–2, double myb34 myb51, myb34 myb122–2, myb51 myb122–2, and triple myb34 myb51 myb122–2 mutants. Three major IGs in shoots (B) and roots (D) are presented. Data are means ± SE from two independent cultivations with four biological replicates (n = 8). Values marked with asterisks are significantly different from Col-0 (Student’s t-test; p < 0.05).
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

5. Figure 4
MYB34, MYB51, and MYB122 Differentially Respond to ABA, JA, ET, SA, and ET/JA Treatments.
Relative expression values of MYB34, MYB51, and MYB122 on MS plates containing: (A) 10 μM abscisic acid (ABA); (B) 50 μM methyl-jasmonate (MeJA); (C) 20 μM 1-aminocyclopropane-1-carboxylic acid (ACC); (D) 200 μM salicylic acid (SA); or (E) 20 μM ACC and 50 μM MeJA. (Expression levels of MYBs in plants grown on MS without supplements = 1.) Data are means ± SE from four independent cultivations with two biological replicates (n = 8). Values marked with asterisks are significantly different from control plants (Student’s t-test; p < 0.05).
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

6. Figure 5
Hormone-Induced Accumulation of IGs Is Affected in myb Loss-of-Function Mutants.
IG content of myb34, myb51, myb122–2, myb34 myb51, myb34 myb51 myb122–2, and wild-type (Col-0) plants grown on MS plates supplemented with: 10 μM ABA (A–C), 50 μM MeJA (D–F), 20 μM ACC (G–I); 200 μM SA (J–L), and 20 μM ACC/ 50 μM MeJA (M–O). The three major IGs I3M (A, D, G, J, M), 4MO-I3M (B, E, H, K, N), and 1MO-I3M (C, F, I, L, O) are presented. Data are presented as means ± SE from three independent cultivations with four biological replicates (n = 12). Values marked with asterisks or circles are significantly different from control plants (* = Col-0 or respective genotype grown on MS = °), respectively (Student’s t-test; p < 0.05).
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

7. Figure 6
MYB122 Is Able to Induce IG Biosynthesis in Leaves of myb34 myb51.
Contents of the three major IGs of 6-week-old myb34 myb51, myb34 myb51 myb122–1/MYB122–1D, and wild-type (Col-0) plants are shown. Data are means ± SE from two independent cultivations with four biological replicates (n = 8). Values marked with asterisks or circles are significantly different from control plants (Col-0) or myb34 myb51, respectively (Student’s t-test; p < 0.05).
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

8. Figure 7
MYB34, MYB51, and MYB122 Reciprocally Regulate Their Expression in Leaves.
Relative expression of MYB34, MYB51, and MYB122 measured in leaves of 6-week-old myb34, myb51, and myb122–2 knockout lines (Col-0 = 1) (A) or MYB34-OX (MYB34/ATR1-1D; Celenza et al., 2005), MYB51-OX (MYB51/HIG1-1D; Gigolashvili et al., 2007a), and MYB122-OX (MYB122-OX; Gigolashvili et al., 2007a) lines (Col-0 = 1) (B). Values in (A) and (B) are means ± SE from three independent cultivations with two biological replicates (n = 6). Values marked with asterisks are significantly (Student’s t-test; p < 0.05) different from control plants (Col-0).
(C) Model for potential activities of MYB34, MYB51, and MYB122 based on the analysis of myb knockout and overexpression mutants. Black arrows show the results obtained from the analysis of myb knockouts; blue arrows show results obtained from the analysis of MYB-overexpressing plants.
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

9. Figure 8 Model for the Regulation of IG Biosynthesis in Arabidopsis.
(A)MYB34, MYB51, and MYB122 are central transcriptional regulators of IG biosynthesis that integrate various signals such as JA, ABA, SA, and ET.
(B, C) Transcription factor TF-1 and TF-2 are as-yet unidentified TFs, which are suggested to be co-expressed with MYB34 and MYB51 and to (co)regulate the biosynthesis of modified IG via the JA/ABA signaling pathway (B) or the SA/ET signaling pathway (C), and to contribute to the production of 1MO-I3M and 4MO-I3M, respectively.
▶ positive regulation/activation; negative regulation/repression; ▶ possible activation; ▶ known interactions in hormone signaling.
Molecular Plant 2014 7, DOI: ( /mp/ssu004)
Copyright © 2014 The Authors. All rights reserved. Terms and Conditions