Working towards recalcitrance mechanisms: increased xylan and homogalacturonan production by overexpression of GAlactUronosylTransferase12 (GAUT12) causes increased recalcitrance and decreased growth in Populus

Abstract

BackgroundThe development of fast-growing hardwood trees as a source of lignocellulosic biomass for biofuel and biomaterial production requires a thorough understanding of the plant cell wall structure and function that underlie the inherent recalcitrance properties of woody biomass. Downregulation of GAUT12.1 in Populus deltoides was recently reported to result in improved biomass saccharification, plant growth, and biomass yield. To further understand GAUT12.1 function in biomass recalcitrance and plant growth, here we report the effects of P. trichocarpa GAUT12.1 overexpression in P. deltoides.ResultsIncreasing GAUT12.1 transcript expression by 7–49% in P. deltoides PtGAUT12.1-overexpression (OE) lines resulted in a nearly complete opposite biomass saccharification and plant growth phenotype to that observed previously in PdGAUT12.1-knockdown (KD) lines. This included significantly reduced glucose, xylose, and total sugar release (12–13%), plant height (6–54%), stem diameter (8–40%), and overall total aerial biomass yield (48–61%) in 3-month-old, greenhouse-grown PtGAUT12.1-OE lines compared to controls. Total lignin content was unaffected by the gene overexpression. Importantly, selected PtGAUT12.1-OE lines retained the recalcitrance and growth phenotypes upon growth for 9 months in the greenhouse and 2.8 years in the field. PtGAUT12.1-OE plants had significantly smaller leaves with lower relative water content, and significantly reduced stem wood xylem cell numbers and size. At the cell wall level, xylose and galacturonic acid contents increased markedly in total cell walls as well as in soluble and insoluble cell wall extracts, consistent with increased amounts of xylan and homogalacturonan in the PtGAUT12.1-OE lines. This led to increased cell wall recalcitrance, as manifested by the 9–15% reduced amounts of recovered extractable wall materials and 8–15% greater amounts of final insoluble pellet in the PtGAUT12.1-OE lines compared to controls.ConclusionsThe combined phenotype and chemotype data from P. deltoides PtGAUT12.1-OE and PdGAUT12.1-KD transgenics clearly establish GAUT12.1 as a recalcitrance- and growth-associated gene in poplar. Overall, the data support the hypothesis that GAUT12.1 synthesizes either an HG-containing primer for xylan synthesis or an HG glycan required for proper xylan deposition, anchoring, and/or architecture in the wall, and the possibility of HG and xylan glycans being connected to each other by a base-sensitive covalent linkage.Electronic supplementary materialThe online version of this article (10.1186/s13068-017-1002-y) contains supplementary material, which is available to authorized users.

Genome-Wide Association Study of Wood Anatomical and Morphological Traits in Populus trichocarpa

“…Another gene with a similar function, Potri.004G111000 (GAUT9) was detected for the same multitrait GWAS from the Corvallis common garden (Chhetri et al, 2019). GAUT8 and GAUT9 belonged to the same clade B1 in a phylogenetic tree based on the GAUT protein family of Arabidopsis thaliana and P. trichocarpa (Biswal et al, 2018). GAUT9 affects leaf size in Populus deltoides (Chhetri et al, 2019) whereas GAUT8 (also known as QUASIMODO1) has high expression in stems in Arabidopsis (Caffall et al, 2009) and is thought to be involved in cell wall pectic homogalacturonan (HG) and xylan biosynthesis (Orfila et al, 2005;Verger et al, 2016).…”

Rhamnogalacturonan‐I is a determinant of cell–cell adhesion in poplar wood

“…Our results are consistent with previous findings that pectin, although present in low abundance in woody tissues, contributes to biomass recalcitrance, as down‐regulation of pectin synthesis enhanced sugar yield in saccharification assays (Biswal et al , ; Biswal et al , ), and over‐expression of GAUT12 to increase HG content decreased saccharification yield (Biswal et al , ). Deletion of a gene cluster encoding a suite of pectolytic enzymes of Caldicellulosiruptor bescii reduced sugar yields in saccharification assays (Chung et al , ), whereas controlled expression of recombinant pectinases enhanced digestibility of A. thaliana (Tomassetti et al , ).…”

“…However, genetic modification of enzymes involved in biosynthesis of the xylan backbone results in decreased stem strength (Brown et al, 2007;Brown et al, 2005;Persson et al, 2007). Over-expression of GALACTURONOSYL TRANSFERASE 12 (GAUT12) increased recalcitrance to enzymatic digestion but also resulted in decreased growth in Populus (Biswal et al, 2018a). Although presumed to be involved in HG backbone synthesis, knockdown of GAUT12 results in reduced xylan content and in increased growth (Biswal et al, 2015).…”

Multitrait genome‐wide association analysis of Populus trichocarpa identifies key polymorphisms controlling morphological and physiological traits

“…Other GAUT genes have also been shown to affect cell wall properties and leaf size in Populus. Downregulation of a GAUT12 homolog in P. deltoides showed decreased xylan and pectin content in the cell wall and increased biomass yield (Biswal et al, 2015), while overexpression showed a reduction in overall plant productivity and resulted in smaller leaves, reduced xylem cell numbers and size, and an increase in the amount of xylose and galacturonic acid in the cell wall (Biswal et al, 2018b). Downregulation of GAUT4 in P. deltoides resulted in decreased pectic homogalacturonan and rhamnogalacturonan II and increased plant height, diameter, leaf area, and biomass (Biswal et al, 2018a).…”