Gene regulatory networks for lignin biosynthesis in switchgrass (Panicum virgatum)

Abstract

Cell wall recalcitrance is the major challenge to improving saccharification efficiency in converting lignocellulose into biofuels. However, information regarding the transcriptional regulation of secondary cell wall biogenesis remains poor in switchgrass (Panicum virgatum), which has been selected as a biofuel crop in the United States. In this study, we present a combination of computational and experimental approaches to develop gene regulatory networks for lignin formation in switchgrass. To screen transcription factors (TFs) involved in lignin biosynthesis, we developed a modified method to perform co-expression network analysis using 14 lignin biosynthesis genes as bait (target) genes. The switchgrass lignin co-expression network was further extended by adding 14 TFs identified in this study, and seven TFs identified in previous studies, as bait genes. Six TFs (PvMYB58/63, PvMYB42/85, PvMYB4, PvWRKY12, PvSND2 and PvSWN2) were targeted to generate overexpressing and/or down-regulated transgenic switchgrass lines. The alteration of lignin content, cell wall composition and/or plant growth in the transgenic plants supported the role of the TFs in controlling secondary wall formation. RNA-seq analysis of four of the transgenic switchgrass lines revealed downstream target genes of the secondary wall-related TFs and crosstalk with other biological pathways. In vitro transactivation assays further confirmed the regulation of specific lignin pathway genes by four of the TFs. Our meta-analysis provides a hierarchical network of TFs and their potential target genes for future manipulation of secondary cell wall formation for lignin modification in switchgrass.

Heteroexpression of Osa-miR319b improved switchgrass biomass yield and feedstock quality by repression of PvPCF5

“…However, we did not detect any β-β bond in tested DELs, which may be due to different lignin extraction and detection methods [35,46]. NAC-MYB-based transcriptional regulatory system regulates secondary cell wall biosynthesis and has been well studied in switchgrass [11,13,47,48]. In Arabidopsis, miR319 target gene TCP4 could directly activate expression of NAC TF AtVND7 (one master regulator in cell wall biosynthesis), which resulted in increases in lignin and cellulose content [33].…”

“…Increasing biomass yield is also accompanied by a higher lignin level, such as ectopic expression ZmGA20ox switchgrass plants [51]. Reducing lignin content by directly down-regulating the lignin biosynthesis enzyme or cell wall-related TFs also resulted in a stunted phenotype [11][12][13]. Fortunately, the negative effect on plant growth may be avoided by regulating some transcription factors that has no direct association with the cell wall biosynthesis.…”

“…7). Two lignin synthesis genes, PvCCR (cinnamoyl CoA reductase) and PvCOMT (caffeic acid O-methyltransferase), showed a similar expression pattern as PvPCF5 [11], showed reduced expression in OE-miR319 compared to WT plants, but had no expression difference in 5sr plants compared to WT plants. The results indicate the reduced lignin content in OE-miR319 and 5sr plants might be…”

“…However, lignocellulose materials are blocked by lignin in the secondary cell walls, resulting in a poor saccharification efficiency [5]. In recent decades, regulating the expression of lignin biosynthesis genes [6][7][8][9][10] or transcription factors (TFs) that involved in secondary cell wall regulation, including members of Liu et al Biotechnol Biofuels (2020) 13:56 MYELOBLASTOSIS (MYB), NAM/ATAF/CUC (NAC), and APETALA2/Ethylene Responsive Factor (AP2/ERF) families [11][12][13][14], could efficiently alter lignin composition and structure and/or secondary cell wall remodeling, leading to enhanced sugar release efficiently [reviewed in 15,16]. However, those improvements were also accompanied with potential side effects such as stunted plant growth [11][12][13], sterile characteristics [12], or sensitivity to biotic and/or abiotic stresses [17,18].…”

Distinct and overlapping functions ofMiscanthus sinensisMYB transcription factors SCM1 and MYB103 in lignin biosynthesis

“…However, in switchgrass expressing PvMYB85A , a close relative of MsSCM1 (Fig. 1), was capable of elevating the S/G ratio (Rao et al ., 2019), whereas MsSCM1 did not change S/G ratio in N. benthamiana (Fig. 4B), highlighting functional nuances between two closely related genes.…”