Dormancy cycling in Arabidopsis seeds is controlled by seasonally distinct hormone-signaling pathways

Abstract

Seeds respond to environmental signals, tuning their dormancy cycles to the seasons and thereby determining the optimum time for plant establishment. The molecular regulation of dormancy cycling is unknown, but an extensive range of mechanisms have been identified in laboratory experiments. Using a targeted investigation of gene expression over the dormancy cycle of Arabidopsis seeds in the field, we investigated how these mechanisms are seasonally coordinated. Depth of dormancy and gene expression patterns were correlated with seasonal changes in soil temperature. The results were consistent with abscisic acid (ABA) signaling linked to deep dormancy in winter being repressed in spring concurrent with enhanced DELLA repression of germination as depth of dormancy decreased. Dormancy increased during winter as soil temperature declined and expression of ABA synthesis (NCED6) and gibberellic acid (GA) catabolism (GA2ox2) genes increased. This was linked to an increase in endogenous ABA that plateaus, but dormancy and DOG1 and MFT expression continued to increase. The expression of SNF1-related protein kinases, SnrK 2.1 and 2.4, also increased consistent with enhanced ABA signaling and sensitivity being modulated by seasonal soil temperature. Dormancy then declined in spring and summer. Endogenous ABA decreased along with positive ABA signaling as expression of ABI2, ABI4, and ABA catabolism (CYP707A2) and GA synthesis (GA3ox1) genes increased. However, during the low-dormancy phase in the summer, expression of transcripts for the germination repressors RGA and RGL2 increased. Unlike deep winter dormancy, this represson can be removed on exposure to light, enabling the completion of germination at the correct time of year.

Fig. 1.

Seasonal changes in dormancy cycling. (A) Changes in dormancy level (AR50) and soil temperature and moisture content measured at seed depth (5 cm) over 12 mo from October 2007. Mean seedling emergence following monthly soil disturbance (n = 4) is also shown. The red horizontal bar is the thermal germination window (Materials and Methods). (B) ABA concentration and changing sensitivity to gibberellic acid (GA) in Cvi seeds recovered from the field. Following recovery, seeds where incubated in the light at 20 °C in the presence of 50–250 μM GA at pH 5.0. (C) Changing thermodormancy in Cvi seeds recovered from the field. Following recovery, seeds where incubated in the light at 5–25 °C. (D) [ABA] and DOG1 expression in relation to dormancy level in the field (AR50 in A). The ABA regression was fitted using the Weibull function (R² = 0.524). The DOG1 regression was fitted using a cubic polynomial function (R² = 0.784). Error bars indicate SEM; n = 3.

Fig. 2.

Gene expression in the deep dormancy phase. (A) Expression of DOG1 (dormancy) and MFT (ABA-induced germination repressor). (B) Expression of two members (SnrK2.1 and -2.4) of the SNF1-related protein kinase subfamily (positive regulators of ABA signaling). (C) Changes in ABI3 (dormancy), NCED6 (ABA biosynthesis), and GA2ox2 (GA catabolism) expression. (D) Expression of FLC (a flowering-time regulator) and ABI5 (ABRE-regulated transcription factor). Shaded area represents the region of deep dormancy. Error bars indicate SEM; n = 3.

Fig. 3.

Gene expression in the shallow dormancy phase. (A) Expression of ABI2 (repressor of ABA signaling), PYL7 (ABA receptor), and CYP707A2 (ABA catabolism) in relation to the changing dormancy level (AR50). (B) Expression of PYR1 (ABA receptor) and ABI4 (control of energy utilization). Shaded area represents the region of shallow dormancy and increasing germination potential. Error bars indicate SEM; n = 3.

Fig. 4.

Gene expression related to germination potential during spatial sensing. (A) Expression of GA3ox1 (GA biosynthesis) and GID1A (GA receptor). (B) Expression of PIL5 and SPT (bHLH transcription factors of the PIF family: germination repressors). (C) Expression of RGA2 and RGL2 (DELLAs: germination repressors). Shaded panel represents the region of increasing germination potential. Error bars indicate SEM; n = 3.

Fig. 5.

Nitrate sensitivity during dormancy cycling in the field. (A) Germination in the light at 20 °C ± 10 mM KNO₃ in relation to changing dormancy level (AR50). (B) Expression of NRT1.1 (nitrate transporter) and NR1 (nitrate reductase). Error bars indicate SEM; n = 3.

Fig. 6.

Seasonal timing of molecular physiological events during dormancy cycling in the field. The height of each bar indicates the amplitude of the response measured across four seasons. Temperature represents the annual fluctuation in soil temperature at seed depth. Depth of dormancy is related to the AR50 of Cvi seeds in the field. Other bars represent seasonal changes in [ABA], gene expression and environmental responses. Temporal sensing represents the slow seasonal change in dormancy for the selection of time of year and climate space (light blue bars). Spatial sensing represents the period of potential rapid change during shallow dormancy if suitable germination conditions are detected (dark blue bars).