Humic Substances Isolated from Recycled Biomass Trigger Jasmonic Acid Biosynthesis and Signalling.

Intensive agriculture maintains high crop yields through chemical inputs, which are well known for their adverse effects on environmental quality and human health. Innovative technologies are required to reduce the risk generated by the extensive and harmful use of pesticides. The plant biostimulants made from humic substances isolated from recyclable biomass offer an alternative approach to address the need for replacing conventional agrochemicals without compromising the crop yield. The stimulatory effects of humic substances are commonly associated with plant hormones, particularly auxins. However, jasmonic acid (JA) is crucial metabolite in mediating the defence responses and governing plant growth and development. This work aimed to evaluate the changes in the biosynthesis and signalling pathway of JA in tomato seedlings treated with humic acids (HA) isolated from vermicompost. We use the tomato model system cultivar Micro-Tom (MT) harbouring a reporter gene fused to a synthetic promoter that responds to jasmonic acid (JERE::GUS). The transcript levels of genes involved in JA generation and activity were also determined using qRT-PCR. The application of HA promoted plant growth and altered the JA status, as revealed by both GUS and qRT-PCR assays. Both JA enzymatic synthesis (LOX, OPR3) and JA signalling genes (JAZ and JAR) were found in higher transcription levels in plants treated with HA. In addition, ethylene (ETR4) and auxin (ARF6) signalling components were positively modulated by HA, revealing a hormonal cross-talk. Our results prove that the plant defence system linked to JA can be emulated by HA application without growth inhibition.

Flagellin and mannitol modulate callose biosynthesis and deposition in soybean seedlings.

Callose is a polymer deposited on the cell wall and is necessary for plant growth and development. Callose is synthesized by genes from the glucan synthase-like family (GSL) and dynamically responds to various types of stress. Callose can inhibit pathogenic infection, in the case of biotic stresses, and maintain cell turgor and stiffen the plant cell wall in abiotic stresses. Here, we report the identification of 23 GSL genes (GmGSL) in the soybean genome. We performed phylogenetic analyses, gene structure prediction, duplication patterns, and expression profiles on several RNA-Seq libraries. Our analyses show that WGD/Segmental duplication contributed to expanding this gene family in soybean. Next, we analyzed the callose responses in soybean under abiotic and biotic stresses. The data show that callose is induced by both osmotic stress and flagellin 22 (flg22) and is related to the activity of ß-1,3-glucanases. By using RT-qPCR, we evaluated the expression of GSL genes during the treatment of soybean roots with mannitol and flg22. The GmGSL23 gene was upregulated in seedlings treated with osmotic stress or flg22, showing the essential role of this gene in the soybean defense response to pathogenic organisms and osmotic stress. Our results provide an important understanding of the role of callose deposition and regulation of GSL genes in response to osmotic stress and flg22 infection in soybean seedlings.

Methyl-CpG binding proteins (MBD) family evolution and conservation in plants.

DNA methylation is an epigenetic mechanism that acts on cytosine residues. The methyl-CpG-binding domain proteins (MBD) are involved in the recognition of methyl-cytosines by activating a signaling cascade that induces the formation of heterochromatin or euchromatin, thereby regulating gene expression. In this study, we analyzed the evolution and conservation of MBD proteins in plants. First, we performed a genome-wide identification and analysis of the MBD family in common bean and soybean, since they have experienced one and two whole-genome duplication events, respectively. We found one pair of MBD paralogs in soybean (GmMBD2) has subfunctionalized after their recent divergence, which was corroborated with their expression profile. Phylogenetic analysis revealed that classes of MBD proteins clustered with human MBD. Interestingly, the MBD9 may have emerged after the hexaploidization event in eudicots. We found that plants and humans share a great similarity in MBDs' binding affinity in the mCpG context. MBD2 and MBD4 from different plant species have the conserved four amino acid residues -Arg (R), Asp (D), Tyr (Y) and Arg (R)- reported to be responsible for MBD-binding in the mCpG. However, MBD8, MBD9, MBD10, and MBD11 underwent substitutions in these residues, suggesting the non-interaction in the mCpG context, but a heterochromatin association as MBD5 and MBD6 from human. This study represents the first genome-wide analysis of the MBD gene family in eurosids I - soybean and common bean. The data presented here contribute towards understanding the evolution of MBDs proteins in plants and their specific binding affinity on mCpG site.

Genome-Wide Analysis of the COBRA-Like Gene Family Supports Gene Expansion through Whole-Genome Duplication in Soybean (Glycine max).

The COBRA-like (COBL) gene family has been associated with the regulation of cell wall expansion and cellulose deposition. COBL mutants result in reduced levels and disorganized deposition of cellulose causing defects in the cell wall and inhibiting plant development. In this study, we report the identification of 24 COBL genes (GmCOBL) in the soybean genome. Phylogenetic analysis revealed that the COBL proteins are divided into two groups, which differ by about 170 amino acids in the N-terminal region. The GmCOBL genes were heterogeneously distributed in 14 of the 20 soybean chromosomes. This study showed that segmental duplication has contributed significantly to the expansion of the COBL family in soybean during all Glycine-specific whole-genome duplication events. The expression profile revealed that the expression of the paralogous genes is highly variable between organs and tissues of the plant. Only 20% of the paralogous gene pairs showed similar expression patterns. The high expression levels of some GmCOBLs suggest they are likely essential for regulating cell expansion during the whole soybean life cycle. Our comprehensive overview of the COBL gene family in soybean provides useful information for further understanding the evolution and diversification of COBL genes in soybean.

Cell wall dynamics and gene expression on soybean embryonic axes during germination.

MAIN CONCLUSION: Identification of the structural changes and cell wall-related genes likely involved in cell wall extension, cellular water balance and cell wall biosynthesis on embryonic axes during germination of soybean seeds. Cell wall is a highly organized and dynamic structure that provides mechanical support for the cell. During seed germination, the cell wall is critical for cell growth and seedling establishment. Although seed germination has been widely studied in several species, key aspects regarding the regulation of cell wall dynamics in germinating embryonic axes remain obscure. Here, we characterize the gene expression patterns of cell wall pathways and investigate their impact on the cell wall dynamics of embryonic axes of germinating soybean seeds. We found 2143 genes involved in cell wall biosynthesis and assembly in the soybean genome. Key cell wall genes were highly expressed at specific germination stages, such as expansins, UDP-Glc epimerases, GT family, cellulose synthases, peroxidases, arabinogalactans, and xyloglucans-related genes. Further, we found that embryonic axes grow through modulation of these specific cell wall genes with no increment in biomass. Cell wall structural analysis revealed a defined pattern of cell expansion and an increase in cellulose content during germination. In addition, we found a clear correlation between these structural changes and expression patterns of cell wall genes during germination. Taken together, our results provide a better understanding of the complex transcriptional regulation of cell wall genes that drive embryonic axes growth and expansion during soybean germination.

A miniature inverted-repeat transposable element, AddIn-MITE, located inside a WD40 gene is conserved in Andropogoneae grasses.

Miniature inverted-repeat transposable elements (MITEs) have been associated with genic regions in plant genomes and may play important roles in the regulation of nearby genes via recruitment of small RNAs (sRNA) to the MITEs loci. We identified eight families of MITEs in the sugarcane genome assembly with MITE-Hunter pipeline. These sequences were found to be upstream, downstream or inserted into 67 genic regions in the genome. The position of the most abundant MITE (Stowaway-like) in genic regions, which we call AddIn-MITE, was confirmed in a WD40 gene. The analysis of four monocot species showed conservation of the AddIn-MITE sequence, with a large number of copies in their genomes. We also investigated the conservation of the AddIn-MITE' position in the WD40 genes from sorghum, maize and, in sugarcane cultivars and wild Saccharum species. In all analyzed plants, AddIn-MITE has located in WD40 intronic region. Furthermore, the role of AddIn-MITE-related sRNA in WD40 genic region was investigated. We found sRNAs preferentially mapped to the AddIn-MITE than to other regions in the WD40 gene in sugarcane. In addition, the analysis of the small RNA distribution patterns in the WD40 gene and the structure of AddIn-MITE, suggests that the MITE region is a proto-miRNA locus in sugarcane. Together, these data provide insights into the AddIn-MITE role in Andropogoneae grasses.