Use of synthetic sugar analogs to probe plant cell wall function in Arabidopsis thaliana
Authors
Villalobos, Jose Alejandro
Issue Date
2020
Type
Dissertation
Language
Keywords
Alternative Title
Abstract
Plants are terrestrial photosynthetic multicellular organisms responsible for producing most of the world’s oxygen. Plants are also responsible for fixating approximately 1011 metric tones of carbon dioxide into reduced forms of carbon that are used for plant metabolism as well as human and animal nutrition. Approximately 70% of plant biomass is derived from plant cell walls. Plant cell walls are polysaccharide-rich extracellular matrices that encapsulate nearly all plant cells and collectively, these polysaccharides are the most abundant biopolymers on the planet. The evolution of cell wall polysaccharides was integral for plants to populate terrestrial environments. Functionally, cell wall polysaccharides are essential for normal plant growth and development, and compromises to the structure or biosynthesis causes severe developmental defects and are often lethal. The essential nature of cell wall polysaccharides creates challenges in conducting genetic studies to elucidate the biosynthesis and function cell wall polysaccharides.The goal of this work was to implement a chemical biology strategy to probe cell wall function in a spatial, temporal, and dose-dependent manner. Our strategy was to use semi-rationally designed monosaccharide analogs that may inhibit glycosyltransferases by competitively binding to their active sites. We screened a small library of monosaccharide analogs and found three analogs: 2-deoxy-2-fluoro-L-fucose (2F-Fuc), 2-deoxy-2-fluoro-D-mannose (2F-Man), and N-dodecyldeoxynojirimycin (ND-DNJ) inhibit growth in Arabidopsis thaliana. Only 2F-Fuc repressed growth by inhibiting fucosylation of a pectic cell wall polysaccharide while 2F-Man and ND-DNJ inhibited growth in unexpected ways. In 2F-Man, the primary mechanism of inhibition was through the glucose repression pathway in Arabidopsis. The mechanism of 2F-Man toxicity was further supported in yeast as it also inhibited growth through a similar glucose repression pathway. Uniquely, ND-DNJ inhibited glycosylation events in sphingolipids in Arabidopsis and as a result inhibited crystalline cellulose deposition, highlighting a novel connection between cellulose and glycosylated sphingolipids. Finally, we discovered suberin deposition occurs in response to cellulose biosynthesis inhibition.