Molecular and functional characterization of sn-glycerol-3-phosphate acyltransferase of plants

  • Author / Creator
    Chen, Xue
  • sn-Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the acylation of sn-1 position of sn-glycerol-3-phosphate to produce lysophosphatidic acid and Coenzyme A. GPATs are involved in several lipid synthetic pathways and play important physiological roles in plant development. The present doctoral thesis includes three related studies, which aim to molecularly and functionally characterize several plant GPAT genes and the encoded enzymes. The first study characterized three endoplasmic reticulum-bound GPAT4s encoded by three homologous GPAT4 genes of Brassica napus (oilseed rape), focusing primarily on their functional divergence and physiological roles in plant development and lipid biosynthesis. The three homologous GPAT4 genes exhibited different expression patterns and altered epigenetic features. Phenotypic rescue of a gpat4 gpat8 Arabidopsis double mutant and analysis of the gpat4 RNAi B. napus lines suggested physiological roles for the GPAT4s in cuticle formation of the rosette leaves, early flower development, pollen development and storage lipid biosynthesis. The second study investigated stable internal reference genes for gene expression studies in B. napus. This project identified four reliable reference genes to be used in gene expression analysis of BnGPAT4 homologues in both vegetative tissues and developing seeds. The third study focused on molecular cloning and biochemical characterization of a soluble plastidial GPAT isolated from a chilling-tolerant plant, western wallflower (Erysimum asperum). A truncated form of recombinant EaGPAT, with the putative transit peptide deleted, was functionally expressed in yeast. A series of enzymatic assays were performed in order to determine the optimum in vitro reaction conditions for the recombinant EaGPAT. The recombinant EaGPAT was further assayed with different acyl-CoAs and exhibited a substrate preference for 18 carbon unsaturated acyl-CoAs. With this substrate preference, the EaGPAT could potentially be used as a biotechnological tool for improving plant chilling-tolerance or increasing unsaturated fatty acid content of seed oil. Overall, the present doctoral studies revealed the functional divergence and important physiological roles of the GPAT4s in B. napus, and biochemically characterized a plastidial GPAT from E. asperum. The knowledge obtained from these studies provides new insights into the role of GPAT in plants and will be useful for further development of biotechnological approaches to modify seed oil biosynthesis in oleaginous crops.

  • Subjects / Keywords
  • Graduation date
    Spring 2011
  • Type of Item
  • Degree
    Doctor of Philosophy
  • DOI
  • License
    This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.