Down-regulation of PvSAMS impairs S-adenosyl-L-methionine and lignin biosynthesis, and improves cell wall digestibility in switchgrass.

S-adenosyl- l-methionine (SAM) is the methyl donor involved in the biosynthesis of guaiacyl (G) and syringyl (S) lignins in vascular plants. SAM is synthesized from methionine through the catalysis of the enzyme S-adenosylmethionine synthase (SAMS). However, the detailed function of SAMS in lignin biosynthesis has not been widely investigated in plants, particularly in monocot species. In this study, we identified PvSAMS genes from switchgrass (Panicum virgatum L.), an important dual-purpose fodder and biofuel crop, and generated numerous transgenic switchgrass lines through PvSAMS RNA interference technology. Down-regulation of PvSAMS reduced the contents of SAM, G-lignins, and S-lignins in the transgenic switchgrass. The methionine and glucoside derivatives of caffeoyl alcohol were found to accumulate in the transgenic plants. Moreover, down-regulation of PvSAMS in switchgrass resulted in brownish stems associated with reduced lignin content and improved cell wall digestibility. Furthermore, transcriptomic analysis revealed that most sulfur deficiency-responsive genes were differentially expressed in the transgenic switchgrass, leading to a significant increase in total sulfur content; thus implying an important role of SAMS in the methionine cycle, lignin biosynthesis, and sulfur assimilation. Taken together, our results suggest that SAMS is a valuable target in lignin manipulation, and that manipulation of PvSAMS can simultaneously regulate the biosynthesis of SAM and methylated monolignols in switchgrass.

Alteration of S-adenosylhomocysteine levels affects lignin biosynthesis in switchgrass.

Methionine (Met) synthesized from aspartate is a fundamental amino acid needed to produce S-adenosylmethionine (SAM) that is an important cofactor for the methylation of monolignols. As a competitive inhibitor of SAM-dependent methylation, the effect of S-adenosylhomocysteine (SAH) on lignin biosynthesis, however, is still largely unknown in plants. Expression levels of Cystathionine γ-synthase (PvCGS) and S-adenosylhomocysteine hydrolase 1 (PvSAHH1) were down-regulated by RNAi technology, respectively, in switchgrass, a dual-purpose forage and biofuel crop. The transgenic switchgrass lines were subjected to studying the impact of SAH on lignin biosynthesis. Our results showed that down-regulation of PvCGS in switchgrass altered the accumulation of aspartate-derived and aromatic amino acids, reduced the content of SAH, enhanced lignin biosynthesis and stunted plant growth. In contrast, down-regulation of PvSAHH1 raised SAH levels in switchgrass, impaired the biosynthesis of both guaiacyl and syringyl lignins and therefore significantly increased saccharification efficiency of cell walls. This work indicates that SAH plays a crucial role in monolignol methylation in switchgrass. Genetic regulation of either PvCGS or PvSAHH1 expression in switchgrass can change intracellular SAH contents and SAM to SAH ratios and therefore affect lignin biosynthesis. Thus, our study suggests that genes involved in Met metabolism are of interest as new valuable targets for cell wall bioengineering in future.

Genome-wide characterization of GRAS family genes in Medicago truncatula reveals their evolutionary dynamics and functional diversification.

The GRAS gene family is a large plant-specific family of transcription factors that are involved in diverse processes during plant development. Medicago truncatula is an ideal model plant for genetic research in legumes, and specifically for studying nodulation, which is crucial for nitrogen fixation. In this study, 59 MtGRAS genes were identified and classified into eight distinct subgroups based on phylogenetic relationships. Motifs located in the C-termini were conserved across the subgroups, while motifs in the N-termini were subfamily specific. Gene duplication was the main evolutionary force for MtGRAS expansion, especially proliferation of the LISCL subgroup. Seventeen duplicated genes showed strong effects of purifying selection and diverse expression patterns, highlighting their functional importance and diversification after duplication. Thirty MtGRAS genes, including NSP1 and NSP2, were preferentially expressed in nodules, indicating possible roles in the process of nodulation. A transcriptome study, combined with gene expression analysis under different stress conditions, suggested potential functions of MtGRAS genes in various biological pathways and stress responses. Taken together, these comprehensive analyses provide basic information for understanding the potential functions of GRAS genes, and will facilitate further discovery of MtGRAS gene functions.

Switchgrass SBP-box transcription factors PvSPL1 and 2 function redundantly to initiate side tillers and affect biomass yield of energy crop.

BACKGROUND: Switchgrass (Panicum virgatum L.) is a dedicated lignocellulosic feedstock for bioenergy production. The SQUAMOSA PROMOTER-BINDING PROTEIN (SBP-box)-LIKE transcription factors (SPLs) change plant architecture and vegetative-to-reproductive phase transition significantly, and as such, they are promising candidates for genetic improvement of switchgrass biomass yield. However, the genome-wide identification and functional characterization of SPL genes have yet to be investigated in herbaceous energy crops. RESULTS: We identified 35 full-length SPL genes in the switchgrass genome. The phylogenetic relationship and expression pattern of PvSPLs provided baseline information for their function characterization. Based on the global overview of PvSPLs, we explored the biological function of miR156-targeted PvSPL1 and PvSPL2, which are closely related members of SPL family in switchgrass. Our results showed that PvSPL1 and PvSPL2 acted redundantly to modulate side tiller initiation, whereas they did not affect phase transition and internode initiation. Consistently, overexpression of the miR156-resistant rPvSPL2 in the miR156-overexpressing transgenic plants greatly reduced tiller initiation, but did not rescue the delayed flowering and increased internode numbers. Furthermore, suppression of PvSPL2 activity in switchgrass increased biomass yield and reduced lignin accumulation, which thereby elevated the total amount of solubilized sugars. CONCLUSIONS: Our results indicate that different miR156-targeted PvSPL subfamily genes function predominantly in certain biological processes in switchgrass. We suggest that PvSPL2 and its paralogs can be utilized as the valuable targets in molecular breeding of energy crops for developing novel germplasms with high biofuel production.