Abstract
Cottonseed oil is recognized as an important oil in food industry for its unique characters: low flavor reversion and the high level of antioxidants (VitaminE) as well as unsaturated fatty acid. However, the cottonseed oil content of cultivated cotton (Gossypium hirsutum) is only around 20%. In this study, we modified the accumulation of oils by the down-regulation of phosphoenolpyruvate carboxylase 1 (GhPEPC1) via RNA interference in transgenic cotton plants. The qRT-PCR and enzyme activity assay revealed that the transcription and expression of GhPEPC1 was dramatically down-regulated in transgenic lines. Consequently, the cottonseed oil content in several transgenic lines showed a significant (P < 0.01) increase (up to 16.7%) without obvious phenotypic changes under filed condition when compared to the control plants. In order to elucidate the molecular mechanism of GhPEPC1 in the regulation of seed oil content, we quantified the expression of the carbon metabolism related genes of transgenic GhPEPC1 RNAi lines by transcriptome analysis. This analysis revealed the decrease of GhPEPC1 expression led to the increase expression of triacylglycerol biosynthesis-related genes, which eventually contributed to the lipid biosynthesis in cotton. This result provides a valuable information for cottonseed oil biosynthesis pathway and shows the potential of creating high cottonseed oil germplasm by RNAi strategy for cotton breeding.
Highlights
Increase in seed oil content was observed from 1% to 4%, mostly at the expense of seed storage protein
Multiple sequence alignment of the PEPC amino acid sequence from G. raimondii genome[5] and G. hirsutum in National Center for Biotechnology Information (NCBI) revealed that G. hirsutum phosphoenolpyruvate carboxylase 1 (GhPEPC1) contains a Ser residue in N terminus, which acts as a phosphorylation site and exclusively belongs to plant PEPCs29–33, and a Ala residue in C3 plant, different from C4 plant (Ser)[32]
Rooted phylogenetic analysis using the PEPCs coding sequence and zea mays (GI: 162461755) acting as outgroup showed two distinct branches, suggesting that GhPEPC1 and GhPEPC2 were belonged to two different branches (Fig. 2a) and might be discrepant in the terms of gene function
Summary
Increase in seed oil content was observed from 1% to 4%, mostly at the expense of seed storage protein. OAA and Pi conjugate with acetyl-CoA to generate citric acid and enter into tricarboxylic acid cycle (TCA) to provide energy as well as other intermediate metabolites for life activities including a variety of intermediates required for the proteins biosynthesis It had been observed in 1989 that the PEPCase activity was positively correlated with protein content and inversely correlated with lipid contents in soybean cultivar[16]. CrPEPC1 over-expression decreased TAG level by 37% and increased PEPC activities by 157–184% These observations suggest that the oil content of algal cells can be controlled by regulation of the CrPEPC1 gene expression[17]. Another important gene in carbon metabolic pathway- Chlamydomonas citrate synthase (CrCIS) was investigated by the same team. Any report about the function of PEPC in cotton carbon flow distribution is not reported
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