Identification and expression analysis of the glycosyltransferase GT43 family members in bamboo reveal their potential function in xylan biosynthesis during rapid growth

Abstract

Background: Xylan is one of the most abundant hemicelluloses and can crosslink cellulose and lignin to increase the stability of cell walls. A number of genes encoding glycosyltransferases play vital roles in xylan biosynthesis in plants, such as those of the GT43 family. However, little is known about glycosyltransferases in bamboo, especially woody bamboo which is a good substitute for timber.

Results: A total of 17 GT43 genes (PeGT43-1 ~ PeGT43-17) were identified in the genome of moso bamboo (Phyllostachys edulis), which belong to three subfamilies with specific motifs. The phylogenetic and collinearity analyses showed that PeGT43s may have undergone gene duplication, as a result of collinearity found in 12 pairs of PeGT43s, and between 17 PeGT43s and 10 OsGT43s. A set of cis-acting elements such as hormones, abiotic stress response and MYB binding elements were found in the promoter of PeGT43s. PeGT43s were expressed differently in 26 tissues, among which the highest expression level was found in the shoots, especially in the rapid elongation zone and nodes. The genes coexpressed with PeGT43s were annotated as associated with polysaccharide metabolism and cell wall biosynthesis. qRT-PCR results showed that the coexpressed genes had similar expression patterns with a significant increase in 4.0 m shoots and a peak in 6.0 m shoots during fast growth. In addition, the xylan content and structural polysaccharide staining intensity in bamboo shoots showed a strong positive correlation with the expression of PeGT43s. Yeast one-hybrid assays demonstrated that PeMYB35 could recognize the 5' UTR/promoter of PeGT43-5 by binding to the SMRE cis-elements.

Conclusions: PeGT43s were found to be adapted to the requirement of xylan biosynthesis during rapid cell elongation and cell wall accumulation, as evidenced by the expression profile of PeGT43s and the rate of xylan accumulation in bamboo shoots. Yeast one-hybrid analysis suggested that PeMYB35 might be involved in xylan biosynthesis by regulating the expression of PeGT43-5 by binding to its 5' UTR/promoter. Our study provides a comprehensive understanding of PeGT43s in moso bamboo and lays a foundation for further functional analysis of PeGT43s for xylan biosynthesis during rapid growth.

Keywords: Glycosyltransferase 43; MYB transcription factors; Phyllostachys edulis; Yeast one-hybrid.

Fig. 1

Phylogenetic tree of GT43 family. Phylogenetic tree of GT43s from Phyllostachys edulis (Pe), Olyra latifolia (Ol), Raddia guianensis (Rg), Guadua angustifolia (Ga), Bonia amplexicaulis (Ba), Brachypodium distachyon (Brandi), Oryza sativa (Os), Zea mays (Zm), Arabidopsis thaliana (At), Gosspium hirsutum (Gorai) and Populus trichocarpa (Potri). Subfamilies of A, B and C are labeled in pink, blue and purple, respectively. PeGT43s and GT43s identified in other species are shaded yellow and pink, respectively; green and red nodes represent monocots and dicots, respectively

Fig. 2

Collinearity of PeGT43s and OsGT43s. The gray lines indicate duplicated blocks, while the blue and red lines indicate the collinear gene pair of PeGT43s and those between PeGT43s and OsGT43s

Fig. 3

Conserved motifs of GT43s. The length of GT43s is presented as black lines. The boxes in different colors distributed on the black line represent the conserved motifs of GT43s

Fig. 4

Cis-acting elements in the promoter of PeGT43s. Upstream sequences (2000 bp) of the PeGT43s start codon were selected as the promoters. A: Corresponding cis-acting elements, the color bar indicates the log₂-based number of these elements. B: SMRE cis-elements, the conserved nucleic acid bases are shaded in grey

Fig. 5

Expression profiles of PeGT43s in different tissues of moso bamboo. The color bar indicates log₂-based fragments per kilobase per million (FPKM). The FPKM value was listed in Table S5

Fig. 6

Expression profiles of PeGT43s in different portions of moso bamboo shoots. PeGT43s with significant transcriptional changes were showed in the folding line chart, using the transcripts per million (TPM) as the value of the vertical coordinate. The TPM value was listed in Table S5

Fig. 7

The co-expression network of PeGT43s

Fig. 8

Relative expression level of PeGT43s analyzed by qRT-PCR. Lowercase letters a, b, c, d and e indicate significant differences in expression level (p < 0.05)

Fig. 9

Relative expression level of co-expressed genes analyzed by qRT-PCR. Lowercase letters a, b, c, d and e indicate significant differences in expression level (p < 0.05)

Fig. 10

Xylan content and structural polysaccharides staining in different height shoots. A: Xylan content. Lowercase letters a, b, c, d and e indicate significant differences in expression level (p < 0.05). B - G: Structural polysaccharide staining intensity of vascular bundle transverse section in 0.5 m (B), 1.0 m (C), 2.0 m (D), 4.0 m (E), 6.0 m (F), 8.0 m (G) shoots. pc: parenchyma cells; sc: sclerenchymatous cell; mv: metaxylem vessel

Fig. 11

Subcellular localization of PeGT43–5 in N. benthamiana leaves expressed with PeGT43–6. A - C: Fluorescent localization of PeGT43–5 and PeGT43–6 fused in-frame to GFP (A), mRFP-tagged Golgi marker (B) and the merged images (C). D: Intensity plot of GFP and mRFP from the transfection shown in (C)

Fig. 12

Y1H assay for the interaction between PeMYB35 and the 5′ UTR/promoter of PeGT43–5. The 5′ UTR/promoter of PeGT43–5 as well as triple SMRE elements were inserted into pHIS2 vector. The reporter and effector constructs were co-transformed into yeast Y187 cells, and the transformed cells were identified by spotting serial dilutions of yeast onto selective medium SD/−Leu/−Trp (DDO) and SD/−Leu/−Trp/−His (TDO) containing 20 mmol·L^− 1 3-amino-1,2,4-triazole (3-AT). P: positive control (p53::HIS2 + pGADT7::53); N: negative control (p53::HIS2 + pGADT7::PeMYB35); Pro: promoter