A Synergistic Genetic Engineering Strategy Induced Triacylglycerol Accumulation in Potato (Solanum tuberosum) Leaf.

Xiao-Yu Xu,Zhongyi Li,Dawar Hussain,Peter J Sharp,Rosangela Devilla,Jiwon Lee,Surinder P Singh,Lauren Venugoban,Qing Liu,Sehrish Akbar,Pushkar Shrestha,Lijun Tian,Melanie Rug,Thomas Vanhercke

doi:10.3389/fpls.2020.00215

Abstract

Potato is the 4th largest staple food in the world currently. As a high biomass crop, potato harbors excellent potential to produce energy-rich compounds such as triacylglycerol as a valuable co-product. We have previously reported that transgenic potato tubers overexpressing WRINKLED1, DIACYLGLYCEROL ACYLTRANSFERASE 1, and OLEOSIN genes produced considerable levels of triacylglycerol. In this study, the same genetic engineering strategy was employed on potato leaves. The overexpression of Arabidopsis thaliana WRINKED1 under the transcriptional control of a senescence-inducible promoter together with Arabidopsis thaliana DIACYLGLYCEROL ACYLTRANSFERASE 1 and Sesamum indicum OLEOSIN driven by the Cauliflower Mosaic Virus 35S promoter and small subunit of Rubisco promoter respectively, resulted in an approximately 30- fold enhancement of triacylglycerols in the senescent transgenic potato leaves compared to the wild type. The increase of triacylglycerol in the transgenic potato leaves was accompanied by perturbations of carbohydrate accumulation, apparent in a reduction in starch content and increased total soluble sugars, as well as changes of polar membrane lipids at different developmental stages. Microscopic and biochemical analysis further indicated that triacylglycerols and lipid droplets could not be produced in chloroplasts, despite the increase and enlargement of plastoglobuli at the senescent stage. Possibly enhanced accumulation of fatty acid phytyl esters in the plastoglobuli were reflected in transgenic potato leaves relative to wild type. It is likely that the plastoglobuli may have hijacked some of the carbon as the result of WRINKED1 expression, which could be a potential factor restricting the effective accumulation of triacylglycerols in potato leaves. Increased lipid production was also observed in potato tubers, which may have affected the tuberization to a certain extent. The expression of transgenes in potato leaf not only altered the carbon partitioning in the photosynthetic source tissue, but also the underground sink organs which highly relies on the leaves in development and energy deposition.

Highlights

Plant vegetative tissues such as leaves are usually viewed as ‘source organs,’ within which a matrix of assimilative photosynthetic activities and metabolite transport proceeds
The transgenic status of these plants was verified by polymerase chain reaction (PCR) of each of the three transgenes being overexpressed, including A. thaliana WRINKLED1 (AtWRI1), A. thaliana diacylglycerol acyltransferase1 (AtDGAT1) and SiOLEOSIN1 from genomic DNA
This study applied the previously reported ‘Push, Pull and Protect’ genetic engineering strategy in potato leaf using the senescence-inducible promoter Senescence Associated Gene 12 (SAG12) to drive the AtWRI1 transcriptional factor, to minimize the undesirable effects of excessive expression of WRI1 on plant development as most of the critical biological processes have been completed at the plant senescent stage (Gregersen et al, 2013; Avila-Ospina et al, 2014; Yang et al, 2015)

Summary

INTRODUCTION

Plant vegetative tissues such as leaves are usually viewed as ‘source organs,’ within which a matrix of assimilative photosynthetic activities and metabolite transport proceeds. We have previously applied the ‘Push, Pull and Protect’ strategy in potato through tuber-specific expression of AtWRI1 driven by the patatin promoter, together with AtDGAT1 and SiOLEOSIN1 which led to an almost 100-fold increase in TAG levels in tuber tissues (Liu et al, 2017a). Earlier attempts failed to enhance TAG accumulation in potato leaf by transforming the ‘Push, Pull, Protect’ construct which was successfully used in generating high oil tobacco leaf (Vanhercke et al, 2014a), likely due to the strong pleiotropic effects of AtWRI1 expression driven by the green tissue active promoter derived from the small subunit of Rubisco (SSU) gene (Qing Liu, unpublished data). The impacts of transgene expression on tuber constituents, morphology and production have been assessed

RESULTS

DISCUSSION

Experimental Procedures

DATA AVAILABILITY STATEMENT