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Characterization of Sphingolipid Biosynthesis and Modification in Plants
Sphingolipids are ubiquitous membrane lipids in eukaryotic cells. Ceramides, the backbone of sphingolipids, are synthesized by ceramide synthases (CerSs), which form amide bonds between long-chain bases (LCBs) and the acyl groups of acyl-CoAs. Class I CerS (“LOH2”) works predominantly towards C16-CoA, whereas Class II CerS (“LOH1/3”) works towards C24-CoA. Plant sphingolipids generally have 16-carbon and 24-carbon as the major acyl groups. However, the sphingolipids of Oryza sativa and Zea mays have 20-carbon and 24-carbon as the predominant acyl groups. This dissertation aims to understand the substrate preference of CerSs in monocots such as rice. Phylogenetic analysis shows the Class I CerSs from grasses form an evolutionarily distinct clade. The CerS I from the rice was expressed in Arabidopsis CerS I mutants, resulting in a GlcCer profile with C20 as the major acyl group. A 16-residue segment was found to determine the C20 acyl chain specificity of the rice CerS I. Plant LCBs generally have two double bonds (d18:2, Δ4, Δ8). However, Arabidopsis leaves have little d18:2. Overexpression of Δ4 desaturase led to an increasing amount of d18:2 but with a corresponding decrease of d18:1 (Δ8). Co-overexpression of both Δ4 desaturase and Class I CerS resulted in increased d18:2 without a reduction in d18:1 (Δ8). The co-overexpression lines showed delayed bolting and yellowish leaves. Sphingolipidome analysis revealed the formation of excessive amounts of ceramides and GlcCers. The impaired phenotype was recovered by overexpressing the GlcCer synthase to channel the excessive ceramides into GlcCers. The core structure of plant GIPCs is composed of an inositol phosphoceramide (IPC) linked to a GlcA. The additional sugar headgroups of GIPCs have not been fully studied in Zea mays. Here, four different GIPC species were identified in Zea mays using mass spectrometry, and the signal intensity/mole factorials for each class were developed. Complex GIPC synthesis is closely related to phosphate and nitrogen metabolism. The content changes of GIPCs were determined from maize grown under nitrogen and phosphate deplete conditions.
Zhang, Dongdong, "Characterization of Sphingolipid Biosynthesis and Modification in Plants" (2021). ETD collection for University of Nebraska - Lincoln. AAI28712988.