Molecular Plant Advance Access published March 7, 2014 Molecular Plant
RESEARCH ARTICLE
Short-Term and Continuing Stresses Differentially Interplay with Multiple Hormones to Regulate Plant Survival and Growth Cangjing Yanga, Jingjing Liua, Xinran Dongb, Zhenying Caia, Weidong Tianb, and Xuelu Wanga,1
ABSTRACT The stress phytohormone, abscisic acid (ABA), plays important roles in facilitating plants to survive and grow well under a wide range of stress conditions. Previous gene expression studies mainly focused on plant responses to short-term ABA treatment, but the effect of sustained ABA treatment and their difference are poorly studied. Here, we treated plants with ABA for 1 h or 9 d, and our genome-wide analysis indicated the differentially regulated genes under the two conditions were tremendously different. We analyzed other hormones’ signaling changes by using their whole sets of known responsive genes as reporters and integrating feedback regulation of their biosynthesis. We found that, under short-term ABA treatment, signaling outputs of growth-promoting hormones, brassinosteroids and gibberellins, and a biotic stress-responsive hormone, jasmonic acid, were significantly inhibited, while auxin and ethylene signaling outputs were promoted. However, sustained ABA treatment repressed cytokinin and gibberellin signaling, but stimulated auxin signaling. Using several sets of hormone-related mutants, we found candidates in corresponding hormonal signaling pathways, including receptors or transcription regulators, are essential in responding to ABA. Our findings indicate interactions of ABA-dependent stress signals with hormones at different levels are involved in plants to survive under transient stress and to adapt to continuing stressful environments. Key words: abscisic acid; crosstalk; short-term stress; continuing stress; phytohormone; survival and growth.
Introduction When encountering unfavorable environments, as sessile organisms, plants initially undergo intensive physiological changes to live, and the survivors need to reprogram their developmental processes to adapt the possibly prolonged hostile environments to efficiently finish their life cycle. Lichtenthaler (1998) proposed that plants’ stress responses can be divided into four phases: response phase (alarm reaction, beginning of stress), restitution phase (stage of resistance, continuing stress), end phase (stage of exhaustion, long-term stress), and regeneration phase. Many previous gene expression profiling studies mainly focused on initial stress responses (response phase) and showed that regulating contents and/or signaling activity of many phytohormones, including abscisic acid (ABA), ethylene (ET), auxin, brassinosteroids (BRs), cytokinins (CKs), gibberellins (GAs), salicylic acid (SA), and jasmonic acid (JA), may be a prevalent strategy for plants to respond to stresses (Cushman and Bohnert, 2000; Kreps et al., 2002; Seki et al., 2002). Under continuing stresses, plants may alter their growth to develop more efficiently.
For example, under continuing drought, plants developed a profounder root system to obtain water from deep soil layers and to enhance water conduction capacity (Levitt, 1980). However, the differential mechanisms of plants at initial stress response phase and continuing stress phase are largely unknown. ABA is a major abiotic stress-responsive hormone (Cutler et al., 2010), whose level is rapidly induced by many abiotic stresses, including salt, drought, and cold (Cutler and Krochko, 1999; Taylor et al., 2000). For instance, accumulated ABA helps plants to survive under drought and salt by enhancing water balance and cellular dehydration tolerance
1 To whom correspondence should be addressed. E-mail [email protected], fax +86-21-6510-2247, tel. +86-21-6510-2247.
© The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS. doi:10.1093/mp/ssu013 Received 6 November 2013; accepted 26 January 2014
Downloaded from http://mplant.oxfordjournals.org/ at The University of Montana on December 15, 2014
a State Key Laboratory of Genetic Engineering, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, 200433, P.R. China b State Key Laboratory of Genetic Engineering, Institute of Biostatistics, School of Life Science, Fudan University, 220 Handan Rd, Shanghai 2004333, P.R. China
Page 2 of 16
Yang et al. • Short-Term and Continuing Stresses Regulate Growth
Taken together, we propose a model to illustrate how shortterm and sustained ABA crosstalk with other hormones, which provides insight into how plants survive and develop at different phases of encountering a stress.
RESULTS Microarray Analysis Revealed a Huge Diversity of Genes Differentially Regulated by Short-Term and Sustained ABA Treatments To understand how plants regulate gene expression in order to survive under short-term stresses and to adapt to continuing unfavorable conditions, we used ABA to mimic ABAdependent stress signals and conducted microarray analysis using RNA samples from 9-day-old seedlings treated with 1 μM ABA for 1 h (as short-term stress samples) or seedlings grown on medium supplemented with 100 nM ABA for 9 d (as continuing stress samples); as the germination rate was too low when grown on 1 μM ABA for 9 d, we used 100 nM ABA to mimic long-term stress, which can significantly inhibit seedling growth (Supplemental Figure 1). An ABA-deficient mutant aba2 was used as the genetic material. We performed transcription profiling using Agilent Arabidopsis Expression gene chips (Redman et al., 2004), and found that 4485 genes were differentially regulated by short-term ABA and 2578 by sustained ABA. Surprisingly, only 1046 genes were in common between the two groups of differently regulated genes (Figure 1A), and 210 of them were regulated in opposite directions, indicating that genes regulated by the two treatments were remarkably different. We then conducted gene ontology (GO) (Ashburner et al., 2000) enrichment analysis, to identify biological processes (BPs) significantly (p 1.4 between ABA and mock treatments. Global coverage for specific subgroup genes is indicated in brackets. (B) Illustration of subgroup genes regulated by short-term and sustained ABA treatments.
Up-regulated under short-term ABA treatment, but Nonchanged under sustained ABA treatment (1572 genes); UD: Up-regulated under short-term ABA treatment, but Downregulated under sustained ABA treatment (118 genes); NU: Non-changed under short-term ABA treatment, but Upregulated under sustained ABA treatment (738 genes); ND: Non-changed under short-term ABA treatment, but Downregulated under sustained ABA treatment (728 genes); DU: Down-regulated under short-term ABA treatment, but Upregulated under sustained ABA treatment (92 genes); DN: Down-regulated under short-term ABA treatment, but Nonchanged under sustained ABA treatment (1516 genes); DD: Down-regulated under both short-term and sustained ABA treatments (387 genes).
Upon short-term ABA treatment, 103 of the 252 BR-upregulated genes (Nemhauser et al., 2006) were significantly altered, and 60.2% (E: 45.0%, E stands for expected ratio of up-regulated to down-regulated genes upon either treatment) of them were down-regulated (p
RESEARCH ARTICLE
Short-Term and Continuing Stresses Differentially Interplay with Multiple Hormones to Regulate Plant Survival and Growth Cangjing Yanga, Jingjing Liua, Xinran Dongb, Zhenying Caia, Weidong Tianb, and Xuelu Wanga,1
ABSTRACT The stress phytohormone, abscisic acid (ABA), plays important roles in facilitating plants to survive and grow well under a wide range of stress conditions. Previous gene expression studies mainly focused on plant responses to short-term ABA treatment, but the effect of sustained ABA treatment and their difference are poorly studied. Here, we treated plants with ABA for 1 h or 9 d, and our genome-wide analysis indicated the differentially regulated genes under the two conditions were tremendously different. We analyzed other hormones’ signaling changes by using their whole sets of known responsive genes as reporters and integrating feedback regulation of their biosynthesis. We found that, under short-term ABA treatment, signaling outputs of growth-promoting hormones, brassinosteroids and gibberellins, and a biotic stress-responsive hormone, jasmonic acid, were significantly inhibited, while auxin and ethylene signaling outputs were promoted. However, sustained ABA treatment repressed cytokinin and gibberellin signaling, but stimulated auxin signaling. Using several sets of hormone-related mutants, we found candidates in corresponding hormonal signaling pathways, including receptors or transcription regulators, are essential in responding to ABA. Our findings indicate interactions of ABA-dependent stress signals with hormones at different levels are involved in plants to survive under transient stress and to adapt to continuing stressful environments. Key words: abscisic acid; crosstalk; short-term stress; continuing stress; phytohormone; survival and growth.
Introduction When encountering unfavorable environments, as sessile organisms, plants initially undergo intensive physiological changes to live, and the survivors need to reprogram their developmental processes to adapt the possibly prolonged hostile environments to efficiently finish their life cycle. Lichtenthaler (1998) proposed that plants’ stress responses can be divided into four phases: response phase (alarm reaction, beginning of stress), restitution phase (stage of resistance, continuing stress), end phase (stage of exhaustion, long-term stress), and regeneration phase. Many previous gene expression profiling studies mainly focused on initial stress responses (response phase) and showed that regulating contents and/or signaling activity of many phytohormones, including abscisic acid (ABA), ethylene (ET), auxin, brassinosteroids (BRs), cytokinins (CKs), gibberellins (GAs), salicylic acid (SA), and jasmonic acid (JA), may be a prevalent strategy for plants to respond to stresses (Cushman and Bohnert, 2000; Kreps et al., 2002; Seki et al., 2002). Under continuing stresses, plants may alter their growth to develop more efficiently.
For example, under continuing drought, plants developed a profounder root system to obtain water from deep soil layers and to enhance water conduction capacity (Levitt, 1980). However, the differential mechanisms of plants at initial stress response phase and continuing stress phase are largely unknown. ABA is a major abiotic stress-responsive hormone (Cutler et al., 2010), whose level is rapidly induced by many abiotic stresses, including salt, drought, and cold (Cutler and Krochko, 1999; Taylor et al., 2000). For instance, accumulated ABA helps plants to survive under drought and salt by enhancing water balance and cellular dehydration tolerance
1 To whom correspondence should be addressed. E-mail [email protected], fax +86-21-6510-2247, tel. +86-21-6510-2247.
© The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS. doi:10.1093/mp/ssu013 Received 6 November 2013; accepted 26 January 2014
Downloaded from http://mplant.oxfordjournals.org/ at The University of Montana on December 15, 2014
a State Key Laboratory of Genetic Engineering, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, 200433, P.R. China b State Key Laboratory of Genetic Engineering, Institute of Biostatistics, School of Life Science, Fudan University, 220 Handan Rd, Shanghai 2004333, P.R. China
Page 2 of 16
Yang et al. • Short-Term and Continuing Stresses Regulate Growth
Taken together, we propose a model to illustrate how shortterm and sustained ABA crosstalk with other hormones, which provides insight into how plants survive and develop at different phases of encountering a stress.
RESULTS Microarray Analysis Revealed a Huge Diversity of Genes Differentially Regulated by Short-Term and Sustained ABA Treatments To understand how plants regulate gene expression in order to survive under short-term stresses and to adapt to continuing unfavorable conditions, we used ABA to mimic ABAdependent stress signals and conducted microarray analysis using RNA samples from 9-day-old seedlings treated with 1 μM ABA for 1 h (as short-term stress samples) or seedlings grown on medium supplemented with 100 nM ABA for 9 d (as continuing stress samples); as the germination rate was too low when grown on 1 μM ABA for 9 d, we used 100 nM ABA to mimic long-term stress, which can significantly inhibit seedling growth (Supplemental Figure 1). An ABA-deficient mutant aba2 was used as the genetic material. We performed transcription profiling using Agilent Arabidopsis Expression gene chips (Redman et al., 2004), and found that 4485 genes were differentially regulated by short-term ABA and 2578 by sustained ABA. Surprisingly, only 1046 genes were in common between the two groups of differently regulated genes (Figure 1A), and 210 of them were regulated in opposite directions, indicating that genes regulated by the two treatments were remarkably different. We then conducted gene ontology (GO) (Ashburner et al., 2000) enrichment analysis, to identify biological processes (BPs) significantly (p 1.4 between ABA and mock treatments. Global coverage for specific subgroup genes is indicated in brackets. (B) Illustration of subgroup genes regulated by short-term and sustained ABA treatments.
Up-regulated under short-term ABA treatment, but Nonchanged under sustained ABA treatment (1572 genes); UD: Up-regulated under short-term ABA treatment, but Downregulated under sustained ABA treatment (118 genes); NU: Non-changed under short-term ABA treatment, but Upregulated under sustained ABA treatment (738 genes); ND: Non-changed under short-term ABA treatment, but Downregulated under sustained ABA treatment (728 genes); DU: Down-regulated under short-term ABA treatment, but Upregulated under sustained ABA treatment (92 genes); DN: Down-regulated under short-term ABA treatment, but Nonchanged under sustained ABA treatment (1516 genes); DD: Down-regulated under both short-term and sustained ABA treatments (387 genes).
Upon short-term ABA treatment, 103 of the 252 BR-upregulated genes (Nemhauser et al., 2006) were significantly altered, and 60.2% (E: 45.0%, E stands for expected ratio of up-regulated to down-regulated genes upon either treatment) of them were down-regulated (p
