Document Type

Dissertation

Degree

Doctor of Philosophy

Major

Biology, Molecular and Cellular Biology

Date of Defense

8-6-2017

Graduate Advisor

Wang, Xuemin

Committee

Ann M. Steffen, Ph.D.

Olivas, Wendy

Kellogg, Elizabeth

Wang, Xuemin

Abstract

Vegetable oils are important commodities as human foods, animal feeds, renewable industrial feedstocks, and biofuels. Many biochemical and regulation events are involved in seed oil formation, including phospholipids metabolism and transcriptional regulation. In this work, I investigated 1) the role of aminoalcoholphosphotransferases (AAPTs) in phospholipid synthesis and plant development, 2) the effects of AAPTs on seed storage lipid production in Arabidopsis and the emerging oil crop Camelina, and 3) the interaction of the lipid mediator phosphatidic acid (PA) with GLABRA2 (GL2), a negative regulator in seed oil production. AAPTs are the enzymes that produce phosphatidylcholine (PC) and phosphatidylethanolamine (PE). The Arabidopsis AAPT1 and AAPT2 are indispensable for plant reproduction and vegetative growth, based on the embryonic lethality phenotype of aapt1/aapt2 double knockout (KO) mutant and the retarded growth in AAPT RNA interference (RNAi) plants. Surprisingly, no change in PC level was observed in any of the mutant lines, while PE level decreased drastically in aapt1 KO, aapt1/aapt1 aapt2/AAPT2 and AAPT RNAi plants. Detailed analysis revealed extensive lipid species changes. The PC species shifted towards C34, while PE species shifted towards C36. The possibility that C34 PC may be exported from chloroplast was not supported by the FA16:0 positional analysis. The direct conversion of PI to PC by base-exchange mechanism was also not detected. Radiolabeling of PC and PE by infiltration of 3H-choline and 3H-ethanolamine suggested AAPT1 has more activity than AAPT2 in leaves, while PC is the preferred product of both AAPTs in vivo. AAPT mutant lines had decreased oil content and increased long chain fatty acids, while AAPT seed specific overexpression (OE) lines had increased oil content. In another work, I found GL2 interacts with PA strongly and specifically. The PA interaction region was mapped to the leucine-zipper region in the DNA-binding domain of GL2. The DNA-binding ability of GL2 was impaired by the interaction with PA. A chromatin immunoprecipitation (ChIP) assay identified some possible GL2 targets and confirmed by gel mobility shift assay. The seed specific RNAi of GL2 increased oil content, but the mucilage in seed coat was not affected.

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