Dual expression of plastidial GPAT1 and LPAT1 regulates triacylglycerol production and the fatty acid profile in Phaeodactylum tricornutum

Abstract

BackgroundMetabolic engineering has emerged as a potential strategy for improving microalgal lipid content through targeted changes to lipid metabolic networks. However, the intricate nature of lipogenesis has impeded metabolic engineering. Therefore, it is very important to identify the crucial metabolic nodes and develop strategies to exploit multiple genes for transgenesis. In an attempt to unravel the microalgal triacylglycerol (TAG) pathway, we overexpressed two key lipogenic genes, glycerol-3-phosphate acyltransferase (GPAT1) and lysophosphatidic acid acyltransferase (LPAT1), in oleaginous Phaeodactylum tricornutum and determined their roles in microalgal lipogenesis.ResultsEngineered P. tricornutum strains showed enhanced growth and photosynthetic efficiency compared with that of the wild-type during the growth phase of the cultivation period. However, both the cell types reached stationary phase on day 7. Overexpression of GPAT1 and LPAT1 increased the TAG content by 2.3-fold under nitrogen-replete conditions without compromising cell growth, and they also orchestrated the expression of other key genes involved in TAG synthesis. The transgenic expression of GPAT1 and LPAT1 influenced the expression of malic enzyme and glucose 6-phosphate dehydrogenase, which enhanced the levels of lipogenic NADPH in the transgenic lines. In addition, GPAT1 and LPAT1 preferred C16 over C18 at the sn-2 position of the glycerol backbone.ConclusionOverexpression of GPAT1 together with LPAT1 significantly enhanced lipid content without affecting growth and photosynthetic efficiency, and they orchestrated the expression of other key photosynthetic and lipogenic genes. The lipid profile for elevated fatty acid content (C16-CoA) demonstrated the involvement of the prokaryotic TAG pathway in marine diatoms. The results suggested that engineering dual metabolic nodes should be possible in microalgal lipid metabolism. This study also provides the first demonstration of the role of the prokaryotic TAG biosynthetic pathway in lipid overproduction and indicates that the fatty acid profile can be tailored to improve lipid production.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1317-3) contains supplementary material, which is available to authorized users.

TAG pathway engineering via GPAT2 concurrently potentiates abiotic stress tolerance and oleaginicity in Phaeodactylum tricornutum

“…In our previous study, we identified GPAT1 in P. tricornutum, which resulted in a significant elevation of neutral lipids by two-fold [6]. Similarly, dual overexpression of GPAT1 and LPAT1 resulted in a 2.3-fold increment in neutral lipids in P. tricornutum [9]. Previous report has shown that each isoform of GPAT exhibit crucial lipogenic role in mammals [27].…”

“…Nevertheless, lipid metabolism in microalgae is complex in nature due to the presence of regulatory factors at various levels, thereby enabling them recalcitrant to metabolic engineering [26]. Consequently, various previous studies have devoted tremendous research efforts to metabolically engineer oleaginous algal species, and studies showed that engineering the TAG pathway has resulted in promising results in microalgae [6,7,9,16].…”

“…Burgeoning investigations and research efforts have offered us an in-depth understanding of the intricate biosynthetic machinery of TAGs in P. tricornutum at genetic level. Recent reports have demonstrated the occurrence of plastidial located TAG biosynthetic pathway in addition to the conventional endoplasmic reticulum localized Kennedy pathway, which advanced our understanding of TAG biogenesis along with fatty acid acylation pattern of the accumulated TAGs [6,7,9]. However, there is still much room for further improvement of microalgal lipogenesis along with the engineered fatty acid profile.…”

Plastidial and ER Triacylglycerol Biosynthesis in a Growth Phase-Dependent Manner in the Heterokont Nannochloropsis oceanica

“…The previous report has shown that overexpression of plastidial LPAAT elevated saturated and polyunsaturated fatty acids, and decreased monounsaturated fatty acids with particular increment and decrement of C16:0 and C16:1 fatty acids, respectively . Similarly, dual overexpression of plastidial GPAT and LPAAT resulted in remarkable lipid enhancement with an altered fatty acid profile in Phaeodactylum tricornutum (Wang et al, 2018). In addition, DGAT, which catalyzes the final committed step of TAG biosynthesis, was remarkably increased during the late growth phase.…”

Development and characterization of a Nannochloropsis mutant with simultaneously enhanced growth and lipid production

“…In this respect, the genetic and metabolic engineering of microalgae are being actively pursued to improve lipid production, which can contribute to reduced biofuel prices [6][7][8][9]. Strain development has focused on the overexpression of metabolic enzymes related to lipid biosynthesis [10][11][12][13][14] and transcriptional regulators [15][16][17][18], or the targeted mutagenesis of competitive pathways of lipid biosynthesis using TALEN, RNAi, and CRISPR/Cas9 techniques [19][20][21][22].…”