A Post-domestication Mutation, Dt2, Triggers Systemic Modification of Divergent and Convergent Pathways Modulating Multiple Agronomic Traits in Soybean.

The semi-determinate stem growth habit in leguminous crops, similar to the "green revolution" semi-dwarf trait in cereals, is a key plant architecture trait that affects several other traits determining grain yield. In soybean semi-determinacy is modulated by a post-domestication gain-of-function mutation in the gene, Dt2, which encodes an MADS-box transcription factor. However, its role in systemic modification of stem growth and other traits is unknown. In this study, we show that Dt2 functions not only as a direct repressor of Dt1, which prevents terminal flowering, but also as a direct activator of putative floral integrator/identity genes including GmSOC1, GmAP1, and GmFUL, which likely promote flowering. We also demonstrate that Dt2 functions as a direct repressor of the putative drought-responsive transcription factor gene GmDREB1D, and as a direct activator of GmSPCH and GmGRP7, which are potentially associated with asymmetric division of young epidermal cells and stomatal opening, respectively, and may affect the plant's water-use efficiency (WUE). Intriguingly, Dt2 was found to be a direct activator or repressor of the precursors of eight microRNAs targeting genes potentially associated with meristem maintenance, flowering time, stomatal density, WUE, and/or stress responses. This study thus reveals the molecular basis of pleiotropy associated with plant productivity, adaptability, and environmental resilience.

Dark period transcriptomic and metabolic profiling of two diverse Eutrema salsugineum accessions.

Eutrema salsugineum is a model species for the study of plant adaptation to abiotic stresses. Two accessions of E. salsugineum, Shandong (SH) and Yukon (YK), exhibit contrasting morphology and biotic and abiotic stress tolerance. Transcriptome profiling and metabolic profiling from tissue samples collected during the dark period were used to investigate the molecular and metabolic bases of these contrasting phenotypes. RNA sequencing identified 17,888 expressed genes, of which 157 were not in the published reference genome, and 65 of which were detected for the first time. Differential expression was detected for only 31 genes. The RNA sequencing data contained 14,808 single nucleotide polymorphisms (SNPs) in transcripts, 3,925 of which are newly identified. Among the differentially expressed genes, there were no obvious candidates for the physiological or morphological differences between SH and YK. Metabolic profiling indicated that YK accumulates free fatty acids and long-chain fatty acid derivatives as compared to SH, whereas sugars are more abundant in SH. Metabolite levels suggest that carbohydrate and respiratory metabolism, including starch degradation, is more active during the first half of the dark period in SH. These metabolic differences may explain the greater biomass accumulation in YK over SH. The accumulation of 56% of the identified metabolites was lower in F1 hybrids than the mid-parent averages and the accumulation of 17% of the metabolites in F1 plants transgressed the level in both parents. Concentrations of several metabolites in F1 hybrids agree with previous studies and suggest a role for primary metabolism in heterosis. The improved annotation of the E. salsugineum genome and newly identified high-quality SNPs will permit accelerated studies using the standing variation in this species to elucidate the mechanisms of its diverse adaptations to the environment.

Growth of Arabidopsis thaliana and Eutrema salsugineum in a closed growing system designed for quantification of plant water use.

The identification of genetic determinants for water-use efficiency (WUE) and their incorporation into crop plants is critical as world water resources are predicted to become less stable over the coming decades. However, quantification of WUE in small model species such as Arabidopsis is difficult because of low plant water loss relative to root zone evaporation. Furthermore, measurements of long-term WUE are labor-intensive and time-consuming. A novel high-throughput closed-container growing system for measuring plant WUE is described. The system eliminates nearly all water loss from the media and does not require irrigation throughout the duration of a typical experiment. Using the model species Arabidopsis thaliana and Eutrema salsugineum, it was confirmed that under growth chamber conditions, this system: (1) eliminates the need for irrigation for as much as 30 days with media water content remaining above 80% full capacity; (2) allows for quantification of WUE in plants with a leaf area as small as ca. 20 cm(2); (3) does not inhibit plant growth; and (4) does not alter media conditions outside of an acceptable range for these species. The growing system provides an efficient high-throughput system for quantifying plant water loss and WUE.

Poplar GTL1 is a Ca2+/calmodulin-binding transcription factor that functions in plant water use efficiency and drought tolerance.

Diminishing global fresh water availability has focused research to elucidate mechanisms of water use in poplar, an economically important species. A GT-2 family trihelix transcription factor that is a determinant of water use efficiency (WUE), PtaGTL1 (GT-2 like 1), was identified in Populus tremula × P. alba (clone 717-IB4). Like other GT-2 family members, PtaGTL1 contains both N- and C-terminal trihelix DNA binding domains. PtaGTL1 expression, driven by the Arabidopsis thaliana AtGTL1 promoter, suppressed the higher WUE and drought tolerance phenotypes of an Arabidopsis GTL1 loss-of-function mutation (gtl1-4). Genetic suppression of gtl1-4 was associated with increased stomatal density due to repression of Arabidopsis STOMATAL DENSITY AND DISTRIBUTION1 (AtSDD1), a negative regulator of stomatal development. Electrophoretic mobility shift assays (EMSA) indicated that a PtaGTL1 C-terminal DNA trihelix binding fragment (PtaGTL1-C) interacted with an AtSDD1 promoter fragment containing the GT3 box (GGTAAA), and this GT3 box was necessary for binding. PtaGTL1-C also interacted with a PtaSDD1 promoter fragment via the GT2 box (GGTAAT). PtaSDD1 encodes a protein with 60% primary sequence identity with AtSDD1. In vitro molecular interaction assays were used to determine that Ca(2+)-loaded calmodulin (CaM) binds to PtaGTL1-C, which was predicted to have a CaM-interaction domain in the first helix of the C-terminal trihelix DNA binding domain. These results indicate that, in Arabidopsis and poplar, GTL1 and SDD1 are fundamental components of stomatal lineage. In addition, PtaGTL1 is a Ca(2+)-CaM binding protein, which infers a mechanism by which environmental stimuli can induce Ca(2+) signatures that would modulate stomatal development and regulate plant water use.

The Arabidopsis GTL1 transcription factor regulates water use efficiency and drought tolerance by modulating stomatal density via transrepression of SDD1.

A goal of modern agriculture is to improve plant drought tolerance and production per amount of water used, referred to as water use efficiency (WUE). Although stomatal density has been linked to WUE, the causal molecular mechanisms have yet to be determined. Arabidopsis thaliana GT-2 LIKE 1 (GTL1) loss-of-function mutations result in increased water deficit tolerance and higher integrated WUE by reducing daytime transpiration without a demonstrable reduction in biomass accumulation. gtl1 plants had higher instantaneous WUE that was attributable to ~25% lower transpiration and stomatal conductance but equivalent CO(2) assimilation. Lower transpiration was associated with higher STOMATAL DENSITY AND DISTRIBUTION1 (SDD1) expression and an ~25% reduction in abaxial stomatal density. GTL1 expression occurred in abaxial epidermal cells where the protein was localized to the nucleus, and its expression was downregulated by water stress. Chromatin immunoprecipitation analysis indicated that GTL1 interacts with a region of the SDD1 promoter that contains a GT3 box. An electrophoretic mobility shift assay was used to determine that the GT3 box is necessary for the interaction between GTL1 and the SDD1 promoter. These results establish that GTL1 negatively regulates WUE by modulating stomatal density via transrepression of SDD1.