Genome-Wide Identification and Characterization of GhCOMT Gene Family during Fiber Development and Verticillium Wilt Resistance in Cotton

Caffeic acid O-methyltransferases (COMTs) play an essential role in lignin synthesis procession, especially in the plant’s phenylalanine metabolic pathway. The content of COMT genes in cotton and the relationship between their expression patterns have not been studied clearly in cotton. In this study, we have identified 190 COMT genes in cotton, which were classified into three groups (I, II and III), and mapped on the cotton chromosomes. In addition, we found that 135 of the 190 COMT genes result from dispersed duplication (DSD) and whole-genome duplication (WGD), indicating that DSD and WGD were the main forces driving COMT gene expansion. The Ka/Ks analysis showed that GhCOMT43 and GhCOMT41 evolved from GaCOMT27 and GrCOMT14 through positive selection. The results of qRT-PCR showed that GhCOMT13, GhCOMT28, GhCOMT39 and GhCOMT55 were related to lignin content during the cotton fiber development. GhCOMT28, GhCOMT39, GhCOMT55, GhCOMT56 and GhCOMT57 responded to Verticillium Wilt (VW) and maybe related to VW resistance through lignin synthesis. Conclusively, this study found that GhCOMTs were highly expressed in the secondary wall thickening stage and VW. These results provide a clue for studying the functions of GhCOMTs in the development of cotton fiber and VW resistance and could lay a foundation for breeding cotton cultivates with higher quantity and high resistance to VW.

Numerical Modeling of Microfluid Dynamics in Xylem Vessels of Khaya grandifoliola

Computational fluid dynamic (CFD) can be used to quantify the internal flow variables of xylem conducting vessels. This study aims to analyze through numerical simulations the xylem water ascent of African mahogany (Khaya grandifoliola) cultivated under different irrigation regimes. We determined a geometric model, defined through the variability of the anatomical structures of the species, observing characteristics of the xylem vessels such as diameter, length, number of pits, and average surface area of the pits. Then we applied numerical simulation through an Eulerian mathematical model with the discretization of volumes via CFD. Compared to other models, we observed that numerical simulation using CFD represented the xylem microstructures in a greater level of detail, contributing to the understanding of the flow of xylem vessels and the interference of its various structures. Analyzing the micrographs, we observed the non-irrigated vessels had a higher number of pits in the secondary wall thickening when compared to the irrigated treatments. This trend influenced the variability of the radial flow of the xylem vessels, causing greater fluid movement in this region and decreasing the influence of the smooth part of the wall, resulting in a lower total resistance of these vessels.

Field evaluation of transgenic hybrid poplars with desirable wood properties and enhanced growth for biofuel production by bicistronic expression of PdGA20ox1 and PtrMYB3 in wood-forming tissue

Background To create an ideotype woody bioenergy crop with desirable growth and biomass properties, we utilized the viral 2A-meidated bicistronic expression strategy to express both PtrMYB3 (MYB46 ortholog of Populus trichocarpa, a master regulator of secondary wall biosynthesis) and PdGA20ox1 (a GA20-oxidase from Pinus densiflora that produces gibberellins) in wood-forming tissue (i.e., developing xylem). Results Transgenic Arabidopsis plants expressing the gene construct DX15::PdGA20ox1-2A-PtrMYB3 showed a significant increase in both stem fresh weight (threefold) and secondary wall thickening (1.27-fold) relative to wild-type (WT) plants. Transgenic poplars harboring the same gene construct grown in a greenhouse for 60 days had a stem fresh weight up to 2.6-fold greater than that of WT plants. In a living modified organism (LMO) field test conducted for 3 months of active growing season, the stem height and diameter growth of the transgenic poplars were 1.7- and 1.6-fold higher than those of WT plants, respectively, with minimal adverse growth defects. Although no significant changes in secondary wall thickening of the stem tissue of the transgenic poplars were observed, cellulose content was increased up to 14.4 wt% compared to WT, resulting in improved saccharification efficiency of the transgenic poplars. Moreover, enhanced woody biomass production by the transgenic poplars was further validated by re-planting in the same LMO field for additional two growing seasons. Conclusions Taken together, these results show considerably enhanced wood formation of our transgenic poplars, with improved wood quality for biofuel production.

Histone methyltransferase ATX1 dynamically regulates fiber secondary cell wall biosynthesis in Arabidopsis inflorescence stem

Secondary wall thickening in the sclerenchyma cells is strictly controlled by a complex network of transcription factors in vascular plants. However, little is known about the epigenetic mechanism regulating secondary wall biosynthesis. In this study, we identified that ARABIDOPSIS HOMOLOG of TRITHORAX1 (ATX1), a H3K4-histone methyltransferase, mediates the regulation of fiber cell wall development in inflorescence stems of Arabidopsis thaliana. Genome-wide analysis revealed that the up-regulation of genes involved in secondary wall formation during stem development is largely coordinated by increasing level of H3K4 tri-methylation. Among all histone methyltransferases for H3K4me3 in Arabidopsis, ATX1 is markedly increased during the inflorescence stem development and loss-of-function mutant atx1 was impaired in secondary wall thickening in interfascicular fibers. Genetic analysis showed that ATX1 positively regulates secondary wall deposition through activating the expression of secondary wall NAC master switch genes, SECONDARY WALL-ASSOCIATED NAC DOMAIN PROTEIN1 (SND1) and NAC SECONDARY WALL THICKENING PROMOTING FACTOR1 (NST1). We further identified that ATX1 directly binds the loci of SND1 and NST1, and activates their expression by increasing H3K4me3 levels at these loci. Taken together, our results reveal that ATX1 plays a key role in the regulation of secondary wall biosynthesis in interfascicular fibers during inflorescence stem development of Arabidopsis.

Computational fluid dynamics model and flow resistance characteristics of Jatropha curcas L xylem vessel

Xylem vessels are the channels used for water transport in Jatropha curcas L. Vessel complexity has a great influence on water transport. Therefore, using anatomical experiments and numerical simulations, the water transport characteristics of J. curcas L xylem vessels with perforation plate and secondary wall thickening (pit structures) were analyzed. The results showed that the xylem vessel in J. curcas provided a low resistance path. The xylem vessel resistance was composed of three elements: smooth vessels, secondary wall thickening and perforation plate. The proportion of smooth vessel resistance was the largest, accounting for 66.20% of the total resistance. Then the secondary wall thickening resistance accounted for 30.20% of the total resistance, and finally the perforation plate resistance accounted for 3.60% of the total resistance. The total resistance of the vessel model was positively correlated with the pit depth, perforation plate height and perforation plate width and negatively correlated with the vessel inner diameter and pit membrane permeability. The vessel inner diameter and the pit depth had a great influence on the total resistance. The total resistance of the vessel inner diameter of 52 µm was 89.15% higher than that of 61 µm, the total resistance of the pit depth of 5.6 µm was 21.98% higher than that of 2.6 µm. The pit structure in the secondary wall thickening caused the vessel to be transported radially, and the radial transmission efficiency of the vessel was positively correlated with the pit depth and pit membrane permeability and negatively correlated with the vessel inner diameter. The pit membrane permeability had the greatest influence on the radial transmission efficiency, and its radial transmission efficiency was 0–5.09%.

Arabidopsis XTH4 and XTH9 contribute to wood cell expansion and secondary wall formation

ABSTRACTIn dicotyledons, xyloglucan is the major hemicellulose of primary walls affecting the load-bearing framework with participation of XTH enzymes. We used loss- and gain-of function approaches to study functions of abundant cambial region expressed XTH4 and XTH9 in secondary growth. In secondarily thickened hypocotyls, these enzymes had positive effects on vessel element expansion and fiber intrusive growth. In addition, they stimulated secondary wall thickening, but reduced secondary xylem production. Cell wall analyses of inflorescence stems revealed changes in lignin, cellulose, and matrix sugar composition, indicating overall increase in secondary versus primary walls in the mutants, indicative of higher xylem production compared to wild type (since secondary walls were thinner). Intriguingly, the number of secondary cell wall layers was increased in xth9 and reduced in xth4, whereas the double mutant xth4x9 displayed intermediate number of layers. These changes correlated with certain Raman signals from the walls, indicating changes in lignin and cellulose. Secondary walls were affected also in the interfascicular fibers where neither XTH4 nor XTH9 were expressed, indicating that these effects were indirect. Transcripts involved in secondary wall biosynthesis and in cell wall integrity sensing, including THE1 and WAK2, were highly induced in the mutants, indicating that deficiency in XTH4 and XTH9 triggers cell wall integrity signaling, which, we propose, stimulates the xylem cell production and modulates secondary wall thickening. Prominent effects of XTH4 and XTH9 on secondary xylem support the hypothesis that altered xyloglucan can affect wood properties both directly and via cell wall integrity sensing.SIGNIFICANCE STATEMENTXyloglucan is a ubiquitous component of primary cell walls in all land plants but has not been so far reported in secondary walls. It is metabolized in muro by cell wall-residing enzymes - xyloglucan endotransglycosylases/hydrolases (XTHs), which are reportedly abundant in vascular tissues, but their role in these tissues is unclear. Here we report that two vascular expressed enzymes in Arabidopsis, XTH4 and XTH9 contribute to the secondary xylem cell radial expansion and intrusive elongation in secondary vascular tissues.Unexpectedly, deficiency in their activities highly affect chemistry and ultrastructure of secondary cell walls by non-cell autonomous mechanisms, including transcriptional induction of secondary wall-related biosynthetic genes and cell wall integrity sensors. These results link xyloglucan metabolism with cell wall integrity pathways, shedding new light on previous reports about prominent effects of xyloglucan metabolism on secondary walls.One sentence summaryXTH4 and XTH9 positively regulate xylem cell expansion and fiber intrusive tip growth, and their deficiency alters secondary wall formation via cell wall integrity sensing mechanisms.

Genetic factor for twisting legume pods identified by fine-mapping of shattering-related traits in azuki bean and yard-long bean

SUMMARYLegumes have evolved a unique manner of seed dispersal in that the seed pods explosively split open with helical tension generated by sclerenchyma on the endocarp. During domestication, azuki bean (Vigna angularis) and yard-long bean (Vigna unguiculata cv-gr. Sesquipedalis) have reduced or lost the sclerenchyma and lost the shattering behavior of seed pods. Here we performed fine-mapping with back-crossed populations and narrowed the candidate genomic region down to 4 kbp in azuki bean and 13 kbp in yard-long bean. Among genes located in these regions, we found MYB26 genes encoded truncated proteins in both the domesticated species. We also found in azuki bean and other legumes that MYB26 is duplicated and only the duplicated copy is expressed in seed pods. Interestingly, in Arabidopsis MYB26 is single copy and is specifically expressed in anther to initiate secondary wall thickening that is required for anther dehiscence. These facts indicated that, in legumes, MYB26 has been duplicated and acquired a new role in development of pod sclerenchyma. However, pod shattering is unfavorable phenotype for harvesting and thus has been selected against by human.

Knockdown of Arabidopsis ROOT UVB SENSITIVE4 Disrupts Anther Dehiscence by Suppressing Secondary Thickening in the Endothecium

ROOT UV-B SENSITIVE4 (RUS4) encodes a protein with no known function that contains a conserved Domain of Unknown Function 647 (DUF647). The DUF647-containing proteins RUS1 and RUS2 have previously been associated with root UV-B-sensing pathway that plays a major role in Arabidopsis early seedling morphogenesis and development. Here, we show that RUS4 knockdown Arabidopsis plants, referred to as amiR-RUS4, were severely reduced in male fertility with indehiscent anthers. Light microscopy of anther sections revealed a significantly reduced secondary wall thickening in the endothecium of amiR-RUS4 anthers. We further show that the transcript abundance of the NAC domain genes NAC SECONDARY WALL THICKENING PROMOTING FACTOR1 (NST1) and NST2, which have been shown to regulate the secondary cell wall thickenings in the anther endothecium, were dramatically reduced in the amiR-RUS4 floral buds. Expression of the secondary cell wall-associated MYB transcription factor genes MYB103 and MYB85 were also strongly reduced in floral buds of the amiR-RUS4 plants. Overexpression of RUS4 led to increased secondary thickening in the endothecium. However, the rus4-2 mutant exhibited no obvious phenotype. Promoter-GUS analysis revealed that the RUS4 promoter was highly active in the anthers, supporting its role in anther development. Taken together, these results suggest that RUS4, probably functions redundantly with other genes, may play an important role in the secondary thickening formation in the anther endothecium by indirectly affecting the expression of secondary cell wall biosynthetic genes.