Volume 6, Issue 2, Pages 369-385 (March 2013) Metabolomic Profiling in Selaginella lepidophylla at Various Hydration States Provides New Insights into the Mechanistic Basis of Desiccation Tolerance  Abou Yobi, Bernard W.M. Wone, Wenxin Xu, Danny C. Alexander, Lining Guo, John A. Ryals, Melvin J. Oliver, John C. Cushman  Molecular Plant  Volume 6, Issue 2, Pages 369-385 (March 2013) DOI: 10.1093/mp/sss155 Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 1 Rehydration/Dehydration Rates in S. lepidophylla. (A) Rehydration time course (h) measured by tracking the percent gain in relative water content (RWC) (blue line and squares) over time and dehydration time course (h) measured by tracking the percent loss in RWC over time (red line and diamonds). Data represent the mean of three independent replicate experiments (n = 9). Error bars represent standard error of the mean. (B) Representative images of S. lepidophylla plants at initial dry state (DRY-1), 50% rehydrated (REH-50), fully rehydrated (HYD), 50% dehydrated (DEH-50), and second dry state (DRY-2). Size bars = 2 cm. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 2 Functional Categorization of 251 Named and Unnamed Metabolites Detected in S. lepidophylla over Five Different RWCs. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 3 Metabolites at Different Hydration States in S. lepidophylla as Defined by Partial Least-Squares-Discriminant Analysis (PLS-DA) Constructed from 204 Metabolites. The plot shows the three components that explains 52.2% of the metabolite signatures (R2 = 0.50; Q2 = 0.40). Upside-down triangles (red) represent initial dry state (DRY-1), squares (brown) represent 50% rehydration (REH-50), asterisks (green) represent fully hydrated (HYD), crosses (cyan) represent 50% dehydrated (DEH-50), and diamonds (moss green) represent the second dry state (DRY-2). Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 4 Metabolites Associated with Energy Metabolism (Glycolysis and TCA Cycle) in S. lepidophylla Profiled during a Rehydration/Dehydration Cycle. Compounds in red indicate greater relative abundance in one or more dry states. Compounds in green indicate greater relative abundance in the hydrated state. Dotted arrows indicate a gap in the pathway and double-headed arrows indicate a reversible reaction. The x-axis represents the percentage of relative water content (% RWC): initial dry state (DRY-1); 50% rehydrated (REH-50); fully rehydrated (HYD); 50% dehydrated (DEH-50); and second dry state (DRY-2). The y-axis box plots indicate the scaled intensity mean (closed triangles) and median (––––) values, top/bottom ranges of boxes indicate upper/lower quartiles, respectively, and top/bottom whiskers indicate the maximum/minimum distribution of the data, respectively. Open circles indicate extreme data points. The p- and q-values for the comparisons are presented in Supplemental Table 1. G-6-P, glucose-6-phosphate; F-6-P, fructose-6-phosphate; M-6-P, mannose-6-phosphate; 3-PG, 3-phosphoglycerate; PEP, phosphoenolpyruvate. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 5 Differences in Sugar Alcohols and Other Sugars in S. lepidophylla Profiled during a Rehydration/Dehydration Cycle. Compounds in green indicate greater relative abundance the hydrated state as shown in box plots. Dotted arrows indicate a gap in the pathway. Box plots are as described in the Figure 4 legend. The p- and q-values for the comparisons are presented in Supplemental Table 1. G-6-P, glucose-6-phosphate; F-6-P, fructose-6-phosphate; Ru5P, ribulose 5-phosphate; Xu5P, xylulose 5-phosphate; E4P, erythrose 4-phosphate; T4P, threose 4-phosphate. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 6 Differences in Amino Acid and Nitrogen Metabolism in S. lepidophylla Profiled during a Rehydration/Dehydration Cycle. Compounds in red indicate greater relative abundance in one or more dry states as shown in box plots. Box plots are as described in the Figure 4 legend. The p- and q-values for the comparisons are presented in Supplemental Table 1. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 7 Differences in γ-Glutamyl Amino Acid and Glutathione Metabolism in S. lepidophylla Profiled during a Rehydration/Dehydration Cycle. Compounds in red or green indicate greater relative abundance in one or more dry states or hydrated state, respectively, as shown in box plots. Dotted arrows indicate a gap in the pathway. Box plots are as described in the Figure 4 legend. The p- and q-values for the comparisons are presented in Supplemental Table 1. Cys-Gly, cysteinylglycine; AA, amino acid; GSH, glutathione (reduced); GSSG, glutathione disulfide (oxidized). Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions

Figure 8 Differences in Nine Unnamed Compounds in S. lepidophylla Profiled during a Rehydration/Dehydration Cycle. Compounds with greater relative abundance in one or more dry states as shown in box plots are presented. Box plots are as described in the Figure 4 legend. The p- and q-values for the comparisons are presented in Supplemental Table 1. Molecular Plant 2013 6, 369-385DOI: (10.1093/mp/sss155) Copyright © 2013 The Authors. All rights reserved. Terms and Conditions