Downregulation of GAUT12 in Populus Deltoides by RNA Silencing Results in Reduced Recalcitrance, Increased Growth and Reduced Xylan and Pectin in a Woody Biofuel Feedstock

Ajaya K. Biswal, Zhangying Hao, Sivakumar Pattathil, Xiaohan Yang, Kim Winkeler, Cassandra Collins, Sushree S. Mohanty, Elizabeth A. Richardson, Ivana Gelineo-Albersheim, Kimberly Hunt, David Ryno, Robert W. Sykes, Geoffrey B. Turner, Angela Ziebell, Erica Gjersing, Wolfgang Lukowitz, Mark F. Davis, Stephen R. Decker, Michael G. Hahn, Debra Mohnen

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Abstract

Background: The inherent recalcitrance of woody bioenergy feedstocks is a major challenge for their use as a source of second-generation biofuel. Secondary cell walls that constitute the majority of hardwood biomass are rich in cellulose, xylan, and lignin. The interactions among these polymers prevent facile accessibility and deconstruction by enzymes and chemicals. Plant biomass that can with minimal pretreatment be degraded into sugars is required to produce renewable biofuels in a cost-effective manner. Results: GAUT12/IRX8 is a putative glycosyltransferase proposed to be involved in secondary cell wall glucuronoxylan and/or pectin biosynthesis based on concomitant reductions of both xylan and the pectin homogalacturonan (HG) in Arabidopsis irx8 mutants. Two GAUT12 homologs exist in Populus trichocarpa, PtGAUT12.1 and PtGAUT12.2. Knockdown expression of both genes simultaneously has been shown to reduce xylan content in Populus wood. We tested the proposition that RNA interference (RNAi) downregulation of GAUT12.1 alone would lead to increased sugar release in Populus wood, that is, reduced recalcitrance, based on the hypothesis that GAUT12 synthesizes a wall structure required for deposition of xylan and that cell walls with less xylan and/or modified cell wall architecture would have reduced recalcitrance. Using an RNAi approach, we generated 11 Populus deltoides transgenic lines with 50 to 67% reduced PdGAUT12.1 transcript expression compared to wild type (WT) and vector controls. Ten of the eleven RNAi lines yielded 4 to 8% greater glucose release upon enzymatic saccharification than the controls. The PdGAUT12.1 knockdown (PdGAUT12.1-KD) lines also displayed 12 to 52% and 12 to 44% increased plant height and radial stem diameter, respectively, compared to the controls. Knockdown of PdGAUT12.1 resulted in a 25 to 47% reduction in galacturonic acid and 17 to 30% reduction in xylose without affecting total lignin content, revealing that in Populus wood as in Arabidopsis, GAUT12 affects both pectin and xylan formation. Analyses of the sugars present in sequential cell wall extracts revealed a reduction of glucuronoxylan and pectic HG and rhamnogalacturonan in extracts from PdGAUT12.1-KD lines. Conclusions: The results show that downregulation of GAUT12.1 leads to a reduction in a population of xylan and pectin during wood formation and to reduced recalcitrance, more easily extractable cell walls, and increased growth in Populus.

Original languageAmerican English
Article number41
Number of pages25
JournalBiotechnology for Biofuels
Volume8
Issue number1
DOIs
StatePublished - 2015

Bibliographical note

Publisher Copyright:
© 2015 Biswal et al.; licensee BioMed Central.

NREL Publication Number

  • NREL/JA-2700-62639

Keywords

  • Biofuel
  • Growth
  • Pectin
  • Populus
  • Saccharification
  • Secondary cell wall
  • Wood development
  • Xylan

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