Identification of genes from the general phenylpropanoid and monolignol-specific metabolism in two sugarcane lignin-contrasting genotypes.

The phenylpropanoid pathway is an important route of secondary metabolism involved in the synthesis of different phenolic compounds such as phenylpropenes, anthocyanins, stilbenoids, flavonoids, and monolignols. The flux toward monolignol biosynthesis through the phenylpropanoid pathway is controlled by specific genes from at least ten families. Lignin polymer is one of the major components of the plant cell wall and is mainly responsible for recalcitrance to saccharification in ethanol production from lignocellulosic biomass. Here, we identified and characterized sugarcane candidate genes from the general phenylpropanoid and monolignol-specific metabolism through a search of the sugarcane EST databases, phylogenetic analysis, a search for conserved amino acid residues important for enzymatic function, and analysis of expression patterns during culm development in two lignin-contrasting genotypes. Of these genes, 15 were cloned and, when available, their loci were identified using the recently released sugarcane genomes from Saccharum hybrid R570 and Saccharum spontaneum cultivars. Our analysis points out that ShPAL1, ShPAL2, ShC4H4, Sh4CL1, ShHCT1, ShC3H1, ShC3H2, ShCCoAOMT1, ShCOMT1, ShF5H1, ShCCR1, ShCAD2, and ShCAD7 are strong candidates to be bona fide lignin biosynthesis genes. Together, the results provide information about the candidate genes involved in monolignol biosynthesis in sugarcane and may provide useful information for further molecular genetic studies in sugarcane.

Silencing CAFFEOYL SHIKIMATE ESTERASE Affects Lignification and Improves Saccharification in Poplar.

Caffeoyl shikimate esterase (CSE) was recently shown to play an essential role in lignin biosynthesis in Arabidopsis (Arabidopsis thaliana) and later in Medicago truncatula However, the general function of this enzyme was recently questioned by the apparent lack of CSE activity in lignifying tissues of different plant species. Here, we show that down-regulation of CSE in hybrid poplar (Populus tremula × Populus alba) resulted in up to 25% reduced lignin deposition, increased levels of p-hydroxyphenyl units in the lignin polymer, and a relatively higher cellulose content. The transgenic trees were morphologically indistinguishable from the wild type. Ultra-high-performance liquid chromatography-mass spectrometry-based phenolic profiling revealed a reduced abundance of several oligolignols containing guaiacyl and syringyl units and their corresponding hydroxycinnamaldehyde units, in agreement with the reduced flux toward coniferyl and sinapyl alcohol. These trees accumulated the CSE substrate caffeoyl shikimate along with other compounds belonging to the metabolic classes of benzenoids and hydroxycinnamates. Furthermore, the reduced lignin amount combined with the relative increase in cellulose content in the CSE down-regulated lines resulted in up to 62% more glucose released per plant upon limited saccharification when no pretreatment was applied and by up to 86% and 91% when acid and alkaline pretreatments were used. Our results show that CSE is not only important for the lignification process in poplar but is also a promising target for the development of improved lignocellulosic biomass crops for sugar platform biorefineries.