BackgroundMicroalgal metabolic engineering holds great promise for the overproduction of a wide range of commercial bioproducts. It demands simultaneous manipulation of multiple metabolic nodes. However, high-efficiency promoters have been lacking.ResultsHere we report a strong constitutive promoter Pt211 in expressing multiple target genes in oleaginous microalga Phaeodactylum tricornutum. Pt211 was revealed to contain significant cis-acting elements. GUS reporter and principal genes glycerol-3-phosphate acyltransferase (GPAT) and diacylglycerol acyltransferase 2 (DGAT2) involved in triacylglycerol biosynthesis were tested under driven of Pt211 in P. tricornutum. GUS staining and qPCR analysis showed strong GUS expression. DGAT2 and GPAT linked with a designed 2A sequence exhibited higher transcript abundances than WT, while algal growth and photosynthesis were not impaired.ConclusionThe total lipid content increased notably by 2.6-fold compared to WT and reached up to 57.5% (dry cell weight). Overall, our findings report a strong promoter and a strategy for coordinated manipulation of complex metabolic pathways.
There has been growing interest in using microalgae as production hosts for a wide range of value‐added compounds. However, microalgal genetic improvement is impeded by lack of genetic tools to concurrently control multiple genes. Here, we identified two novel strong promoters, designated Pt202 and Pt667, and delineated their potential role on simultaneously driving the expression of key lipogenic genes in Phaeodactylum tricornutum. In silico analyses of the identified promoter sequences predicted the presence of essential core cis elements such as TATA and CAAT boxes. Regulatory role of the promoters was preliminarily assessed by using GUS reporter which demonstrated strong GUS expression. Thereafter, two key lipogenic genes including malic enzyme (PtME) and 5‐desaturase (PtD5b), were overexpressed by the two promoters Pt202 and Pt667, respectively, in P. tricornutum. Combinatorial gene overexpression did not impair general physiological performance, meanwhile neutral lipid content was remarkably increased by 2.4‐fold. GC‐MS analysis of fatty acid methyl esters revealed that eicosapentaenoic acid (EPA; C20:5) was increased significantly. The findings augment a crucial kit to microalgal genetic tools that could facilitate the multiple‐gene expression driven by various promoters, and promote microalgae for industrial bioproduction.
Burgeoning commercial applications of catechol have led to its excessive accumulation in the environment, thereby posing a severe ecological threat. Bioremediation has emerged as a promising solution. The potential of the microalga Crypthecodinium cohnii to degrade catechol and use the byproduct as a carbon source was investigated in this study. Catechol significantly increased C. cohnii growth and was rapidly catabolized within 60 h of cultivation. Transcriptomic analysis highlighted the key genes involved in catechol degradation. Real-time polymerase chain reaction (RT-PCR) analysis showed that transcription of key genes CatA, CatB, and SaID involved in the ortho-cleavage pathway was remarkably increased by 2.9-, 4.2-, and 2.4-fold, respectively. Key primary metabolite content was also markedly altered, with a specific increment in polyunsaturated fatty acids. Electron microscopy and antioxidant analysis showed that C. cohnii could tolerate catechol treatment without morphological aberrations or oxidative stress. The findings provide a strategy for C. cohnii in the bioremediation of catechol and concurrent polyunsaturated fatty acids (PUFA) accumulation.
Background
The development of affordable strategy to concurrently enhance microalgal biomass and biocomponents is warranted for commercial applications. Here, we investigated the efficacy of a cheap and natural polyphenol, tannic acid, on regulating metabolic and physiological properties of oleaginous microalga Phaeodactylum tricornutum to overproduce biocompounds.
Result
Tannic acid provision regulated the key metabolic pathways to enhance algal biomass and lipids in a dose-dependent manner without direct impact on photosynthesis. It reduced oxidative stress and reallocated carbon precursors towards lipogenesis. Lipidomic analyses showed that tannic acid provision unprecedentedly regulated the key lipogenic pathways, enhanced glyco- and neutral-lipids by 1.29- and 1.54-fold, respectively, whereas phospholipids were significantly altered. Tannic acid facilitated polyunsaturated fatty acid overproduction with a specific increment of EPA and DHA by 1.18- and 2.25-fold, respectively. Transcriptomic analysis demonstrated that tannic acid upregulated the expression of multiple genes involved in lipogenesis.
Conclusion
Here, we dissected the potential of tannic acid, a natural and cheaper polyphenol, on concurrently enhancing lipids and PUFAs without impairing physiological properties in P. tricornutum. The findings provide novel insights into the mechanistic roles of polyphenol as a potential chemical modulator.
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