Splice Variants of the Castor WRI1 Gene Upregulate Fatty Acid and Oil Biosynthesis When Expressed in Tobacco Leaves.

Xia-Jie Ji,Jin-Ai Xue,Xue Mao,Bao-Ling Liu,Qing-Ting Hao,Run-Zhi Li

doi:10.3390/ijms19010146

Xia-Jie Ji, Jin-Ai Xue + Show 4 more

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https://doi.org/10.3390/ijms19010146

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Abstract

The plant-specific WRINKLED1 (WRI1) is a member of the AP2/EREBP class of transcription factors that positively regulate oil biosynthesis in plant tissues. Limited information is available for the role of WRI1 in oil biosynthesis in castor bean (Ricinus connunis L.), an important industrial oil crop. Here, we report the identification of two alternatively spliced transcripts of RcWRI1, designated as RcWRI1-A and RcWRI1-B. The open reading frames of RcWRI1-A (1341 bp) and RcWRI1-B (1332 bp) differ by a stretch of 9 bp, such that the predicted RcWRI1-B lacks the three amino acid residues “VYL” that are present in RcWRI1-A. The RcWRI1-A transcript is present in flowers, leaves, pericarps and developing seeds, while the RcWRI1-B mRNA is only detectable in developing seeds. When the two isoforms were individually introduced into an Arabidopsis wri1-1 loss-of-function mutant, total fatty acid content was almost restored to the wild-type level, and the percentage of the wrinkled seeds was largely reduced in the transgenic lines relative to the wri1-1 mutant line. Transient expression of each RcWRI1 splice isoform in N. benthamiana leaves upregulated the expression of the WRI1 target genes, and consequently increased the oil content by 4.3–4.9 fold when compared with the controls, and RcWRI1-B appeared to be more active than RcWRI1-A. Both RcWRI1-A and RcWRI1-B can be used as a key transcriptional regulator to enhance fatty acid and oil biosynthesis in leafy biomass.

Highlights

Vegetable oils stored in plant seeds are predominantly composed of triacylglycerols (TAGs), glycerol esters with three fatty acids that serve as an energy reserve for catabolism during germination [1]
A primer pair was designed based on the open reading frame (ORF) of this gene, which was subsequently used to amplify the cDNA encoding RcWRI1 by RT-PCR
Sequence analysis revealed that the long3foofr1m4 (1341 bp) of RcWRI1 was nearly identical with the short form (1332 bp) of RcWRI1, with the short lfaocrkminlgacaksitnrgetcahstorfe9tc-bhponf u9-cblepotnidueclseeonticdoedsinegnc“oVdYinLg” “(FVigYuLr”e (1F)i.gTuhree l1o)n. gThanedlosnhgoratncdDsNhAorstocfDRNcWAsRIo1f wRceWreRnIa1mweedreRncWamReI1d-ARcaWndRIR1c-AWRanI1d-BR,crWesRpIe1c-tBiv, erelys.pectively

Summary

Introduction

Vegetable oils stored in plant seeds are predominantly composed of triacylglycerols (TAGs), glycerol esters with three fatty acids that serve as an energy reserve for catabolism during germination [1]. Understanding the molecular and biochemical mechanisms underlying fatty acid (FA)/oil biosynthesis and regulation is important to undertake this endeavor. Current knowledge of the FA and TAG biosynthesis has been mainly obtained from the study of the model plant Arabidopsis [4,5]. These studies have revealed a series of enzymes that are involved in converting photosynthate sucrose to TAG [6,7,8]. The resulting TAGs within the ER membrane are budded off as specialized structures called oil bodies surrounded by a single-layer phospholipids and proteins

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