<p></p><p></p><p>Abiotic stresses, such as cold, are serious agricultural problems resulting in substantial crop and revenue losses. Soybean (<i>Glycine max</i>) is an important worldwide crop for food, feed, fuel, and other products. Soybean has long been considered to be cold-intolerant and incapable of cold acclimation. In contrast to these reports, this study demonstrates that cold acclimation improved freezing tolerance in the domestic soybean cultivar ‘Williams 82’ with 50% enhancement of freezing tolerance after 5.2 +\- 0.6 days of cold exposure. Decreases in light dependent photosynthetic function and efficiency accompanied cold treatment. These decreases were due to an increase in photon dissipation likely driven by a decrease in plastoquinone (PQ) pool size limiting electron flow from photosystem II (PSII) to photosystem I (PSI). Cold-induced damage to operational photosynthesis began at 25 minutes of cold exposure and maximal photosynthesis was disrupted after 6 to 7 hours of cold exposure. Cold exposure caused severe photodamage leading to the loss of PSII reaction centers and photosynthetic efficiency.</p> <p>Comparisons of eight cultivars of <i>G. max</i> demonstrated a weak correlation between cold acclimation and northern cultivars versus southern cultivars. In the non-domesticated soybean species <i>Glycine soja</i>, the germination rate after cold imbibition was positively correlated with seedling cold acclimation potential. However, the overall cold acclimation potential in <i>G. soja</i> was equal to that of domestic soybean <i>G. max</i> reducing the enthusiasm for the “wild” soybean as an additional source of genetic diversity for cold tolerance. </p> <p> </p> <p>Despite being relatively cold intolerant, the soybean genome possesses homologs of the major cold responsive CBF/DREB1 transcription factors. These genes are cold-induced in soybean in a similar pattern to that of the cold tolerant model plant species Arabidopsis thaliana. In Arabidopsis, EIN3, a major component of the ethylene signaling pathway, is a negative transcriptional regulator of CBF/DREB1. In contrast to <i>AtEIN3</i> transcript levels which do not change during cold treatment in Arabidopsis, we observed a cold-dependent 3.6 fold increase in <i>GmEIN3 </i>transcript levels in soybean. We hypothesized that this increase could prevent effective CBF/DREB1 cold regulation in soybean. Analysis of our newly developed cold responsive reporter (<i>AtRD29Aprom::GFP/GUS</i>) soybean transgenic lines demonstrated that inhibition of the ethylene pathway via foliar sprays (AVG, 1-MCP, and silver nitrate) resulted in significant cold-induced GUS activity. Transcripts of <i>GmEIN3A;1</i> increased in response to ethylene pathway stimulation (ACC and ethephon) and decreased in response to ethylene pathway inhibition in the cold. Additionally, in the cold, inhibition of the ethylene pathway resulted in a significant increase in transcripts of <i>GmDREB1A;1</i> and <i>GmDREB1A;2</i> and stimulation of the ethylene pathway led to a decrease in <i>GmDREB1A;1</i> and <i>GmDREB1B;1</i> transcripts. To assess the physiological effects of these transcriptional changes; electrolyte leakage, lipid oxidation, free proline content, and photosynthesis were examined. Improvement in electrolyte leakage, a measure of freezing tolerance, was seen only under silver nitrate treatment. Only 1-MCP treatment resulted in significantly decreased lipid oxidation. Transcripts for CBF/DREB1 downstream targets (containing the consensus CRT/DRE motifs) significantly decreased in plants treated with ethylene pathway stimulators in the cold; however, ethylene pathway inhibition generally produced no increase over basal cold levels. </p> <p> </p> <p>To identify if GmEIN3A;1 was capable of binding to <i>GmDREB1</i> promoters, the negative regulator GmEIN3A;1 and the positive regulator GmICE1A were cloned and expressed in Escherichia coli (E. coli). Preliminary binding results indicated that GmEIN3A;1 can bind to a double stranded section of the GmDREB1A;1 promoter containing putative EIN3 and ICE1 binding sites. GmICE1A is capable of binding to the same section of the <i>GmDREB1A;1</i> promoter, though only when single stranded. Additional experiments will be required to demonstrate that GmEIN3A;1 and GmICE1A are capable of binding to the <i>GmDREB1A;1</i> promoter and this work provides the tools to answer these questions. </p> <p> </p> <p>Overall, this work provides evidence that the ethylene pathway transcriptionally inhibits the CBF/DREB1 pathway in soybean through the action of GmEIN3A;1. Yet when <i>GmCBF/DREB1</i> transcripts are upregulated by ethylene pathway inhibition, no consistent change in downstream targets was observed. These data indicate that the limitation in cold tolerance in soybean is due to a yet unidentified target downstream of CBF/DREB1 transcription.</p><p></p><p></p>

Bread wheat (Triticum aestivum L.) is one of the most important cereal crops grown in the world. It has great importance because it constitutes a major source of carbohydrate for more than one third of the world’s population (Budak et al., 2013). In the last three decades, drought conditions are becoming more widespread in wheat production areas including Europe, Australia and Asia, and it is considered a major cause of reduced wheat growth and productivity in most developing countries with semi-arid climates. Drought constitutes the most important threat for wheat production in Iraq and especially for the Kurdistan Regional Governate owing to the limited source of water during at least some part of the growing period. Because wheat is considered a staple food and has economic importance for the Kurdistan Regional Government research is needed to determine the production capacity of Iraqi wheat varieties under drought stress and the potential for the maximization of the drought tolerance response. The soil moisture holding capacity of the intended growing medium was measured gravimetrically in pots with and without wheat plants and correlated with the soil capacitance measured using a TDR Theta Probe (Delta-T Devices). This was used to determine the available water content of the soil (AWC) and to control and manage the watering regimes during drought studies. The results of a study of the response of different cultivars of Iraqi wheat (Triticum aestivum L.) to watering regimes of 70% and 50% showed that drought stress had a significant effect on the biomass and yield traits especially tiller number and stem bundle weight compared to normal conditions. The highest significant difference was observed for cv. Tamooz 2 in comparison to Adana 99, but there was a little difference between cvs. Rizgary and Sham 6. The effect of the exogenous application of salicylic acid (SA) and molybdenum (Mo) on drought tolerance of cvs. Tamooz 2 and Adana 99, showed that Tamooz 2 had higher values for growth characteristics and higher yield potential when sprayed with a lower concentration of SA (1.44 mM) under well-watered conditions in comparison with Adana 99. The effect of spraying variety Tamooz 2 with SA at different growth stages indicated that biomass production and yield components (the number of spikes/pot, grain dry weight and average 1000 grain dry weight) significantly increased at both stem+flower as well as leaf+stem+flower sprayings for plants subjected to drought. Also, SA treatments at stem extension and flowering had a positive effect on the up-regulation of the drought response gene CBF/DREB under drought stress conditions. These findings indicate that agronomic treatments with exogenous applications of salicylic acid and molybdenum could help to reduce the effects of drought in the field.

Indiana University-Purdue University Indianapolis (IUPUI) / Abiotic stresses, such as cold, are serious agricultural problems resulting in substantial crop and revenue losses. Soybean (Glycine max) is an important worldwide crop for food, feed, fuel, and other products. Soybean has long been considered to be cold-intolerant and incapable of cold acclimation. In contrast to these reports, this study demonstrates that cold acclimation improved freezing tolerance in the domestic soybean cultivar ‘Williams 82’ with 50% enhancement of freezing tolerance after 5.2 +\- 0.6 days of cold exposure. Decreases in light dependent photosynthetic function and efficiency accompanied cold treatment. These decreases were due to an increase in photon dissipation likely driven by a decrease in plastoquinone (PQ) pool size limiting electron flow from photosystem II (PSII) to photosystem I (PSI). Cold-induced damage to operational photosynthesis began at 25 minutes of cold exposure and maximal photosynthesis was disrupted after 6 to 7 hours of cold exposure. Cold exposure caused severe photodamage leading to the loss of PSII reaction centers and photosynthetic efficiency. Comparisons of eight cultivars of G. max demonstrated a weak correlation between cold acclimation and northern cultivars versus southern cultivars. In the non-domesticated soybean species Glycine soja, the germination rate after cold imbibition was positively correlated with seedling cold acclimation potential. However, the overall cold acclimation potential in G. soja was equal to that of domestic soybean G. max reducing the enthusiasm for the “wild” soybean as an additional source of genetic diversity for cold tolerance. Despite being relatively cold intolerant, the soybean genome possesses homologs of the major cold responsive CBF/DREB1 transcription factors. These genes are cold-induced in soybean in a similar pattern to that of the cold tolerant model plant species Arabidopsis thaliana. In Arabidopsis, EIN3, a major component of the ethylene signaling pathway, is a negative transcriptional regulator of CBF/DREB1. In contrast to AtEIN3 transcript levels which do not change during cold treatment in Arabidopsis, we observed a cold-dependent 3.6 fold increase in GmEIN3 transcript levels in soybean. We hypothesized that this increase could prevent effective CBF/DREB1 cold regulation in soybean. Analysis of our newly developed cold responsive reporter (AtRD29Aprom::GFP/GUS) soybean transgenic lines demonstrated that inhibition of the ethylene pathway via foliar sprays (AVG, 1-MCP, and silver nitrate) resulted in significant cold-induced GUS activity. Transcripts of GmEIN3A;1 increased in response to ethylene pathway stimulation (ACC and ethephon) and decreased in response to ethylene pathway inhibition in the cold. Additionally, in the cold, inhibition of the ethylene pathway resulted in a significant increase in transcripts of GmDREB1A;1 and GmDREB1A;2 and stimulation of the ethylene pathway led to a decrease in GmDREB1A;1 and GmDREB1B;1 transcripts. To assess the physiological effects of these transcriptional changes; electrolyte leakage, lipid oxidation, free proline content, and photosynthesis were examined. Improvement in electrolyte leakage, a measure of freezing tolerance, was seen only under silver nitrate treatment. Only 1-MCP treatment resulted in significantly decreased lipid oxidation. Transcripts for CBF/DREB1 downstream targets (containing the consensus CRT/DRE motifs) significantly decreased in plants treated with ethylene pathway stimulators in the cold; however, ethylene pathway inhibition generally produced no increase over basal cold levels. To identify if GmEIN3A;1 was capable of binding to GmDREB1 promoters, the negative regulator GmEIN3A;1 and the positive regulator GmICE1A were cloned and expressed in Escherichia coli (E. coli). Preliminary binding results indicated that GmEIN3A;1 can bind to a double stranded section of the GmDREB1A;1 promoter containing putative EIN3 and ICE1 binding sites. GmICE1A is capable of binding to the same section of the GmDREB1A;1 promoter, though only when single stranded. Additional experiments will be required to demonstrate that GmEIN3A;1 and GmICE1A are capable of binding to the GmDREB1A;1 promoter and this work provides the tools to answer these questions. Overall, this work provides evidence that the ethylene pathway transcriptionally inhibits the CBF/DREB1 pathway in soybean through the action of GmEIN3A;1. Yet when GmCBF/DREB1 transcripts are upregulated by ethylene pathway inhibition, no consistent change in downstream targets was observed. These data indicate that the limitation in cold tolerance in soybean is due to a yet unidentified target downstream of CBF/DREB1 transcription.