Brassinosteroids affect wood development and properties of Fraxinus mandshurica

IntroductionXylem development plays a crucial role in wood formation in woody plants. In recent years, there has been growing attention towards the impact of brassinosteroids (BRs) on this xylem development. In the present study, we evaluated the dynamic variation of xylem development in Fraxinus mandshurica (female parent, M8) and a novel interspecific hybrid F. mandshurica × Fraxinus sogdiana (1601) from May to August 2020.MethodsWe obtained RNA-Seq transcriptomes of three tissue types (xylem, phloem, and leaf) to identify the differences in xylem-differentially expressed genes (X-DEGs) and xylem-specifically expressed genes (X-SEGs) in M8 and 1601 variants. We then further evaluated these genes via weighted gene co-expression network analysis (WGCNA) alongside overexpressing FmCPD, a BR biosynthesis enzyme gene, in transient transgenic F. mandshurica.ResultsOur results indicated that the xylem development cycle of 1601 was extended by 2 weeks compared to that of M8. In addition, during the later wood development stages (secondary wall thickening) of 1601, an increased cellulose content (14%) and a reduced lignin content (11%) was observed. Furthermore, vessel length and width increased by 67% and 37%, respectively, in 1601 compared with those of M8. A total of 4589 X-DEGs were identified, including enzymes related to phenylpropane metabolism, galactose metabolism, BR synthesis, and signal transduction pathways. WGCNA identified hub X-SEGs involved in cellulose synthesis and BR signaling in the 1601 wood formation–related module (CESA8, COR1, C3H14, and C3H15); in contrast, genes involved in phenylpropane metabolism were significantly enriched in the M8 wood formation–related module (CCoAOMT and CCR). Moreover, overexpression of FmCPD in transient transgenic F. mandshurica affected the expression of genes associated with lignin and cellulose biosynthesis signal transduction. Finally, BR content was determined to be approximately 20% lower in the M8 xylem than in the 1601 xylem, and the exogenous application of BRs (24-epi brassinolide) significantly increased the number of xylem cell layers and altered the composition of the secondary cell walls in F. mandshurica.DiscussionOur findings suggest that BR biosynthesis and signaling play a critical role in the differing wood development and properties observed between M8 and 1601 F. mandshurica

Characterization of endophytic fungi from Acer ginnala Maxim. in an artificial plantation: media effect and tissue-dependent variation.

The community of endophytic fungi associated with Acer ginnala, a common tree in northeastern China, was investigated. Four media, PDA, Czapek's, WA and Sabouraud's, were used to inoculate explants from seeds, annual twigs and perennial twigs (xylem and bark). Media strongly affected the isolated species number, but not colonization frequency (CF) or isolation frequency (IF). To investigate media effect further, a Principal Component Analysis (PCA) was done. As a result, two components accounted for 86.502% of the total variance were extracted. These two components were named as PDA-determined factor (accounted for 45.139% of the total variance) and Czapek's-determined factor (accounted for 41.363% of the total variance), respectively. This result suggested that only two media, PDA and Czapek's, could be used instead of all four media in this study without affecting the isolation results significantly. In total, ten taxa were isolated in this study. Alternaria sp., Phomopsis sp., Neurospora sp. and Phoma sp. were dominant endophytes while Pleosporales Incertae Sedis sp., Cladosporium sp., Trichoderma sp. and Epicoccum sp. were rare taxa. Different tissues/organs had different endophyte assemblages. All tissue/organ pairs had low Bray-Curtis indices (<0.3) except for bark and annual twigs (0.63). Compared to perennial twigs, annual twigs had a lower taxon number, lower isolate number, lower endophyte dominance and diversity indices. Seeds had distinct assemblage, lower similarity and similar low diversity indices to annual twigs. These results suggested that tissue type determines the endophyte assemblage while age determines the diversity

Characteristics and Expression Analysis of <i>FmTCP15</i> under Abiotic Stresses and Hormones and Interact with DELLA Protein in <i>Fraxinus</i> <i>mandshurica</i> Rupr.

The TEOSINTE BRANCHED1, CYCLOIDEA, and PROLIFERATION CELL FACTOR (TCP) transcription factor is a plant-specific gene family and acts on multiple functional genes in controlling growth, development, stress response, and the circadian clock. In this study, a class I member of the TCP family from Fraxinus mandshurica Rupr. was isolated and named FmTCP15, which encoded a protein of 362 amino acids. Protein structures were analyzed and five ligand binding sites were predicted. The phylogenetic relationship showed that FmTCP15 was most closely related to Solanaceae and Plantaginaceae. FmTCP15 was localized in the nuclei of F. mandshurica protoplast cells and highly expressed in cotyledons. The expression pattern revealed the FmTCP15 response to multiple abiotic stresses and hormone signals. Downstream genes for transient overexpression of FmTCP15 in seedlings were also investigated. A yeast two-hybrid assay confirmed that FmTCP15 could interact with DELLA proteins. FmTCP15 participated in the GA-signaling pathway, responded to abiotic stresses and hormone signals, and regulated multiple genes in these biological processes. Our study revealed the potential value of FmTCP15 for understanding the molecular mechanisms of stress and hormone signal responses

Putrescine Promotes Betulin Accumulation in Suspension Cell Cultures of Betula platyphylla by Regulating NO and NH4+ Production

Putrescine (Put) can enhance secondary metabolite production, but its intrinsic regulatory mechanism remains unclear. In this study, Put treatment promoted betulin production and gene expression of lupeol synthase (LUS), one of betulin synthetic enzymes. The maximum betulin content and gene expression level of LUS was 4.25 mg&middot;g&minus;1 DW and 8.25 at 12 h after 1 mmol&middot;L&minus;1 Put treatment, approximately two- and four-times that in the control, respectively. Put treatment increased the content of nitric oxide (NO) and its biosynthetic enzyme activity of nitrate reductase (NR) and NO synthase (NOS). Pretreatment of the birch suspension cells with NO-specific scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline- 1-oxyl-3-oxide (cPTIO), NR inhibitor sodium azide (NaN3), and NOS inhibitor NG-nitro-L-Arg methyl ester (L-NAME) decreased Put-triggered NO generation and blocked Put-induced betulin production. Put treatment improved the content of NH4+ and its assimilation enzyme activity of glutamate synthase and glutamate dehydrogenase. NH4+ supplementation also promoted NO and betulin production. Thus, the above data indicated that Put-induced NO was essential for betulin production. NO derived from NR, NOS, and NH4+ mediated betulin production in birch suspension cell cultures under Put treatment

Correspondence analysis of endophytes and tissues.

<p>The legends see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0046785#pone-0046785-g002" target="_blank">Figure 2</a>.</p

Principal Component Analysis on the effect of media.

<p>Alt: <i>Alternaria</i> sp.; Pen: <i>Penicillium</i> sp.; Neu: <i>Neurospora</i> sp.; Cla: <i>Cladosporium</i> sp.; Phoma: <i>Phoma</i> sp.; Fus: <i>Fusarium</i> sp.; Phomo: <i>Pomopsis</i> sp.; Non: Pleosporales <i>Incertae Sedis</i> sp.; Tri: <i>Trichoderma</i> sp.; Epc: <i>Epicoccum</i> sp. Loadings over 0.7 were in bold.</p>*<p>:percentage of variance.</p>#<p>:Cumulative percentage.</p

Relative frequency of fungal endophytes from <i>A. ginnala</i>.

<p>Relative frequency (RF) means isolate frequency of a given endophyte divided by the sum of isolate frequencies of all endophytes×100.</p

Media effect on species isolated from <i>A. ginnala.</i>

<p>Alt: <i>Alternaria</i> sp.; Pen: <i>Penicillium</i> sp.; Neu: <i>Neurospora</i> sp.; Cla: <i>Cladosporium</i> sp.; Phoma: <i>Phoma</i> sp.; Fus: <i>Fusarium</i> sp.; Phomo: <i>Phomopsis</i> sp.; Non: Pleosporales <i>Incertae Sedis</i> sp.; Tri: <i>Trichoderma</i> sp.; Epc: <i>Epicoccum</i> sp.</p

Colonization and isolation frequency of fungal endophytes from <i>A. ginnala.</i>

<p>Colonization and isolation frequency of fungal endophytes from <i>A. ginnala.</i></p