The cuticular wax serves as the outermost hydrophobic barrier of plants against nonstomatal water loss and various environmental stresses. An objective of this study was to investigate the contribution of the mutualistic fungal endophyte Epichloë gansuensis to leaf cuticular wax of Achnatherum inebrians under different soil moisture availability. Through a pot experiment and gas chromatography−mass spectrometry (GC−MS) analysis, our results indicated that the hydrocarbons were the dominant components of leaf cuticular wax, and the proportion of alcohols, aldehydes, amines, and ethers varied with the presence or absence of E. gansuensis and different soil moisture availability. Amines and ethers are unique in endophyte-free (EF) A. inebrians plants and endophyte-infected (EI) A. inebrians plants, respectively. By transcriptome analysis, we found a total of 13 differentially expressed genes (DEGs) related to cuticular biosynthesis, including FabG, desB, SSI2, fadD, BiP, KCS, KAR, FAR, and ABCB1. A model is proposed which provides insights for understanding cuticular wax biosynthesis in the association of A. inebrians plants with E. gansuensis. These results may help guide the functional analyses of candidate genes important for improving the protective layer of cuticular wax of endophyte-symbiotic plants.
Alfalfa (Medicago sativa) is an important food and feed crop which rich in mineral sources. The WUSCHEL-related homeobox (WOX) gene family plays important roles in plant development and identification of putative gene families, their structure, and potential functions is a primary step for not only understanding the genetic mechanisms behind various biological process but also for genetic improvement. A variety of computational tools, including MAFFT, HMMER, hidden Markov models, Pfam, SMART, MEGA, ProtTest, BLASTn, and BRAD, among others, were used. We identified 34 MsWOX genes based on a systematic analysis of the alfalfa plant genome spread in eight chromosomes. This is an expansion of the gene family which we attribute to observed chromosomal duplications. Sequence alignment analysis revealed 61 conserved proteins containing a homeodomain. Phylogenetic study sung reveal five evolutionary clades with 15 motif distributions. Gene structure analysis reveals various exon, intron, and untranslated structures which are consistent in genes from similar clades. Functional analysis prediction of promoter regions reveals various transcription binding sites containing key growth, development, and stress-responsive transcription factor families such as MYB, ERF, AP2, and NAC which are spread across the genes. Most of the genes are predicted to be in the nucleus. Also, there are duplication events in some genes which explain the expansion of the family. The present research provides a clue on the potential roles of MsWOX family genes that will be useful for further understanding their functional roles in alfalfa plants.
Soil salinity imposes a major threat to plant growth and agricultural productivity. Despite being one of the most common fodder crops in saline locations, alfalfa is vulnerable to salt stress. Jasmonic acid (JA) is a phytohormone that influences plant response to abiotic stimuli such as salt stress. However, key genes and pathways by which JA-mediated salt tolerance of alfalfa are little known. A comprehensive transcriptome analysis was performed to elucidate the underlying molecular mechanisms of JA-mediated salt tolerance. The transcripts regulated by salt (S) compared to control (C) and JA+salt (JS) compared to C were investigated. Venn diagram and expression pattern of DEGs indicated that JS further altered a series of genes expression regulated by salt treatment, implying the roles of JA in priming salt tolerance. Enrichment analysis revealed that DEGs exclusively regulated by JS treatment belonged to primary or secondary metabolism, respiratory electron transport chain, and oxidative stress resistance. Alternatively, splicing (AS) was induced by salt alone or JA combined treatment, with skipped exon (SE) events predominately. DEGs undergo exon skipping involving some enriched items mentioned above and transcription factors. Finally, the gene expressions were validated using quantitative polymerase chain reaction (qPCR), which produced results that agreed with the sequencing results. Taken together, these findings suggest that JA modulates the expression of genes related to energy supply and antioxidant capacity at both the transcriptional and post-transcriptional levels, possibly through the involvement of transcription factors and AS events.
Cuticular wax plays a critical role as a plant protectant against various environmental stresses. We predicted that the presence of the mutualistic fungal endophyte Epichloë gansuensis in Achnatherum inebrians would change both the composition of leaf cuticular wax as plants aged during the growing season and the gene expression levels associated with the wax biosynthesis pathway. Endophyte-infected (EI) and endophyte-free (EF) A. inebrians plants were established for a four-month pot experiment. In agreement with our prediction, the presence of E. gansuensis can change the composition of leaf cuticular wax at different growing periods, particularly the proportion of esters, fatty acids and hydrocarbons. The proportion of fatty acids in EI plants was lower than that in EF plants. The proportion of hydrocarbons increased and esters decreased as plants grew. Furthermore, we found 11 DEGs coding for proteins involved in cuticular wax biosynthesis, including FabF, FAB2, ECR, FAR, CER1, ABCB1 and SEC61A. The present study highlights the significant contribution of E. gansuensis to leaf cuticular wax composition and biosynthesis in A. inebrians plants.
Agarwood, the resin part of Aquilaria spp., is valued in medicine, perfumes, and incense. The most important components of agarwood are sesquiterpenes, which are produced only when a healthy tree is wounded. Agarwood sesquiterpene synthase 1 (ASS1) is one of key enzymes responsible for the biosynthesis of sesquiterpenes in Aquilaria sinensis (Lour.) Gilg, and it is a typical wound-inducible synthase. To elucidate its regulatory mechanism at the transcriptional level, a 978-bp sequence upstream of the translation initiation codon ATG of the promoter for ASS1 was cloned. Computational analysis revealed that this promoter contained many known cis-elements including several defense related transcriptional factor-binding boxes. To functionally validate the promoter, a 5' truncated fragment fused with the β-glucuronidase (GUS) reporter gene was used for generating stable transgenic Arabidopsis plants. The spatial and temporal expression patterns of GUS in transgenic Arabidopsis showed that the promoter of ASS1 was induced by mechanical wound and mainly expressed in vascular bundles. Subcellular localization showed that ASS1 localized in the nucleus and plasma membrane. Here, identification of the ASS1 promoter not only lays a foundation for studying its transcriptional regulation, but also provides clues for studying the synthesis mechanism of agarwood sesquiterpenes.
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