Functional genomics and systems genetics of cellulose biosynthesis in Eucalyptus

The globally emerging bioeconomy demands rapid advancement in the sustainable production and utilization of bio-based raw materials for a multitude of downstream applications, particularly in the areas of food, health and bioenergy and biomaterials. These needs, particularly pertaining to plant productivity, quality and stress tolerance, will need to be addressed with advanced biotechnology strategies, which accelerate progress beyond what has been achieved with traditional breeding and cultivation methods. Woody biomass is a readily available source of renewable carbon, and trees from the genus Eucalyptus, displaying superior growth and wood properties and established agricultural practices worldwide, are attractive candidates as short-rotation (5-9 years) feedstocks for biofuels and biomaterials. Guiding advanced strategies in biotechnology in Eucalyptus and other biomass feedstocks requires a sophisticated understanding of the molecular underpinnings of carbon allocation and cell wall biology. In the work presented here, we aimed to characterize the molecular biology of cellulose biosynthesis in Eucalyptus xylem (developing wood) and identify genes, processes and pathways that are linked to and possibly influence this process. We achieved this by detailed characterization of field-grown Eucalyptus hybrid trees, utilizing RNA-sequencing technology and metabolomics of xylem as well as measuring wood properties that are thought to impact the efficiency of industrial processing. Given the lack of information with regards to gene expression in Eucalyptus trees, a major aim was to characterize transcriptomes from various tissues and organs, including a cellulose-enriched form of xylem called tension wood. This involved challenging bioinformatics, which resulted in a high quality assembly and publication of a comprehensive gene catalogue for Eucalyptus, which was one of the first short-read RNA-sequencing based de novo assembly from a eukaryotic organism. We also characterized and modelled the properties of cellulose and xylan biosynthetic pathways as a biological system, the parts of which are segregating in Eucalyptus hybrid tree populations, which has generated novel insights into the allocation and partitioning of sequestered carbon between cellulose, xylan and lignin during active secondary cell wall deposition in woody stem tissues. This research has made important contributions to the field of Eucalyptus biology, but also to the broader field of secondary cell wall biosynthesis in plants, specifically providing (i) resources for transcriptome analysis in a large woody perennial (ii) new biological insight into carbon allocation for polysaccharide biosynthesis in wood, and (iii) annotation and discovery of candidate genes and pathways that may influence wood chemical composition and structures. Importantly, we find that cellulose and xylan biosynthetic genes are transcriptionally hardwired in their co-regulation (along with other important processes for cellulose and xylan transport and deposition), likely due to the fact that they utilize a common source of sucrose-derived carbon for cell wall biosynthesis and the production of sufficient energy to do so. This co-regulation appears to be distinct from the regulation of other cell wall biopolymers. Furthermore, evidence from xylem gene expression and metabolite availability in xylem, as research has made important contributions to the field of Eucalyptus biology, but also to the broader field of secondary cell wall biosynthesis in plants, specifically providing (i) resources for transcriptome analysis in a large woody perennial (ii) new biological insight into carbon allocation for polysaccharide biosynthesis in wood, and (iii) annotation and discovery of candidate genes and pathways that may influence wood chemical composition and structures. Importantly, we find that cellulose and xylan biosynthetic genes are transcriptionally hardwired in their co-regulation (along with other important processes for cellulose and xylan transport and deposition), likely due to the fact that they utilize a common source of sucrose-derived carbon for cell wall biosynthesis and the production of sufficient energy to do so. This co-regulation appears to be distinct from the regulation of other cell wall biopolymers. Furthermore, evidence from xylem gene expression and metabolite availability in xylem, as well as from wood properties of field-grown trees, supports a model in which sucrose-derived cytosolic fructose is shunted to the production of lignin precursors during cellulose and xylan biosynthesis. This model parsimoniously explains a mechanism for trees to partition carbon between polysaccharide and lignin synthesis, and provides exciting new questions and potential strategies to influence carbon allocation in the secondary cell walls of woody plants.Thesis (PhD)--University of Pretoria, 2013.Wood and Fibre Molecular Genetics (WFMG) ProgrammeNational Research Foundation (NRF)Technology and Human Resources for Industry Programme (THRIP)GeneticsPhDUnrestricte