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Abiotic stress, such as drought, osmotic, and salinity stresses, seriously affects plant growth and crop production. Studying stress-resistant genes that enhance plant stress tolerance is an efficient way to facilitate the breeding of crop species with high stress tolerance. In this study, we reported that the core circadian clock component, the LATE ELONGATED HYPOCOTYL (LHY) orthologue MtLHY, plays a positive role in salt stress response in Medicago truncatula. The expression of MtLHY was induced by salt stress, and loss-of-function mutants of MtLHY were shown to be hypersensitive to salt treatment. However, overexpression of MtLHY improved salt stress tolerance through a higher accumulation of flavonoids. Consistently, exogenous flavonol application improved the salt stress tolerance in M. truncatula. Additionally, MtLHY was identified as a transcriptional activator of the flavonol synthase gene, MtFLS. Our findings revealed that MtLHY confers plant salt stress tolerance, at least by modulating the flavonoid biosynthesis pathway, which provides insight into salt stress tolerance that links the circadian clock with flavonoid biosynthesis.

The molecular mechanisms underlying the diversity of leaf shapes have been of great interest to researchers. Leaf shape depends on the pattern of serrations and the degree of indentation of leaf margins. Multiple transcription factors and hormone signaling pathways are involved in this process. In this study, we characterized the developmental roles of SMALL AND SERRATED LEAF (SSL) by analyzing a recessive mutant in the model legume Medicago truncatula. An ortholog of Arabidopsis thaliana GA3-oxidase 1 (GA3ox1), MtGA3ox1/SSL, is required for GA biosynthesis. Loss of function in MtGA3ox1 results in the small plant and lateral organs. The prominent phenotype of the mtga3ox1 mutant is a more pronounced leaf margin, indicating the critical role of GA level in leaf margin formation. Moreover, 35S:MtDELLA2ΔDELLA and 35S:MtARF3 transgenic plants display leaves with a deeply wavy margin, which resembles those of mtga3ox1. Further investigations show that MtGA3ox1 is under the control of MtDELLA1/2/3–MtGAF1 complex-dependent feedback regulation. Further, MtARF3 behaves as a competitive inhibitor of MtDELLA2/3–MtGAF1 complexes to repress the expression of MtGA3ox1 indirectly. These findings suggest that GA feedback regulatory circuits play a fundamental role in leaf margin formation, in which the posttranslational interaction between transcription factors functions as an additional feature.

As sessile organisms, plants perceive, respond, and adapt to the environmental changes for optimal growth and survival. The plant growth and fitness are enhanced by circadian clocks through coordination of numerous biological events. In legume species, nitrogen-fixing root nodules were developed as the plant organs specialized for symbiotic transfer of nitrogen between microsymbiont and host. Here, we report that the endogenous circadian rhythm in nodules is regulated by MtLHY in legume species Medicago truncatula. Loss of function of MtLHY leads to a reduction in the number of nodules formed, resulting in a diminished ability to assimilate nitrogen. The operation of the 24-h rhythm in shoot is further influenced by the availability of nitrogen produced by the nodules, leading to the irregulated nyctinastic leaf movement and reduced biomass in mtlhy mutants. These data shed new light on the roles of MtLHY in the orchestration of circadian oscillator in nodules and shoots, which provides a mechanistic link between nodulation, nitrogen assimilation, and clock function.

Plant leaves have evolved into diverse shapes and LATE MERISTEM IDENTITY1 (LMI1) and its putative paralogous genes encode homeodomain leucine zipper transcription factors that are proposed evolutionary hotspots for the regulation of leaf development in plants. However, the LMI1-mediated regulatory mechanism underlying leaf shape formation is largely unknown. MtLMI1a and MtLMI1b are putative orthologs of LMI1 in the model legume barrelclover (Medicago truncatula). Here, we investigated the role of MtLMI1a and MtLMI1b in leaf margin morphogenesis by characterizing loss-of-function mutants. MtLMI1a and MtLMI1b are expressed along leaf margin in a near-complementary pattern, and they redundantly promote development of leaf margin serrations, as revealed by the relatively smooth leaf margin in their double mutants. Moreover, MtLMI1s directly activate expression of SMOOTH LEAF MARGIN1 (SLM1), which encodes an auxin efflux carrier, thereby regulating auxin distribution along the leaf margin. Further analysis indicates that MtLMI1s genetically interact with NO APICAL MERISTEM (MtNAM) and the ARGONAUTE7 (MtAGO7)-mediated trans-acting short interfering RNA3 (TAS3 ta-siRNA) pathway to develop the final leaf margin shape. The participation of MtLMI1s in auxin-dependent leaf margin formation is interesting in the context of functional conservation. Furthermore, the diverse expression patterns of LMI1s and their putative paralogs within key domains are important drivers for functional specialization, despite their functional equivalency among species.

Analyzing the mixtures obtained from hydrogenation or hydrogenolysis reactions of biomass-derived platform chemicals is challenging work. With the development and improvement of NMR techniques, the NMR spectrometer proves to be an alternative and highly efficient piece of equipment for the rapid analysis of complex mixtures without the need for tedious purification. Herein, diffusion-ordered spectroscopy (DOSY) is applied in analyzing four model mixtures, which consist of the reactants and products from hydrogenation/hydrogenolysis reactions of biomass-derived platform chemicals. The results show that the DOSY technique can pseudoseparate most components in the model mixtures. The 1D selective gradient TOCSY technique is used as a supporting tool in the cases where the DOSY technique cannot clearly distinguish between the components of the mixtures. This is generally a problem when components in the mixture have very similar diffusion coefficients or severe overlap of peaks. The results show that DOSY and...