Research progress and prospects in lipid metabolic engineering of eukaryotic microalgae

doi:10.12211/2096-8280.2023-044

Abstract

Abstract:

Microalgae represent a diverse group of photosynthetic organisms that are widely found in various ecosystems on the Earth. They play a crucial role in carbon dioxide bio-fixation. Apart from their efficient growth through photosynthesis, many microalgae can also grow robustly under heterotrophic and mixotrophic conditions for high biomass production. Due to their high lipid content and the presence of diverse fatty acid and lipid species, microalgae have a wide range of applications in industries of energy, chemicals, and food. However, the high production cost associated with microalgae-based bioenergy poses a significant challenge for large-scale implementation. To overcome this, there is a growing interest in engineering microalgae to enhance lipid biosynthesis and accumulation, which holds promise for improving the economic feasibility of microalgal lipid production. This requires a better understanding of lipid metabolism and regulation in microalgae. This article provides an overview of recent advances in the elucidation of lipid metabolic pathways, the roles of key enzyme genes involved in lipid metabolism, and the transcriptional regulation of lipid metabolic pathways under different cultivation conditions in eukaryotic microalgae. It also summarizes strategies for metabolic engineering aiming for manipulating lipid biosynthesis-related enzymes, transcription factors, and competing pathways to increase lipid content and/or modify fatty acid composition in microalgae. Integrated analysis of genomics, transcriptomics, and proteomics data can help identify crucial nodes and key regulators in lipid metabolism, facilitating the identification of potential targets for metabolic engineering. Furthermore, the rapid development of genetic tools and gene editing technologies has significantly improved transformation efficiency and enabled precise gene modification, providing a foundation for genetic engineering of microalgae. By reshaping energy and carbon metabolic pathways, it becomes possible to design and optimize lipid biosynthesis processes in microalgae for a better production. Further research and exploration in genetic tools, gene editing technologies, metabolic pathway regulation, and large-scale implementation are of utmost importance for driving the research and development of microalgal lipid engineering.

Key words: eukaryotic microalgae, biorefineries, lipid metabolism, energy metabolism, synthetic biology

摘要：

真核微藻作为一类重要的生物资源，脂质含量高，广泛应用于能源、化工和食品等领域。然而，真核微藻生物能源成本偏高，其产业化应用仍然面临着一系列挑战。通过代谢工程手段改造微藻，促进脂质的合成与积累，可提高微藻脂质生产的经济可行性。本文介绍真核微藻脂质代谢途径和关键酶基因，并总结了不同培养条件下代谢途径相关基因在转录水平上的变化。还探讨通过代谢工程调控脂质合成相关酶、转录因子和竞争途径等方法，以提高微藻脂质含量和调整脂肪酸组成。通过基因组学、转录组学和蛋白质组学数据的整合分析可揭示脂质代谢中的关键节点和主效调控因子，有助于确定代谢工程的潜在目标。此外，基因工具和基因编辑技术的开发和拓展可显著提高转化效率，实现对微藻底盘细胞的精准改造。通过重塑能量和碳代谢途径，可设计优化微藻脂质生物合成过程。在微藻遗传工具、基因编辑技术、代谢通路调控和产业化等方面的进一步研究和探索对于推动微藻脂质工程的研究和发展具有重要意义。

关键词: 真核微藻, 生物质精制, 脂质代谢, 能量代谢, 合成生物学

CLC Number:

Q819

Han SUN, Jin LIU. Research progress and prospects in lipid metabolic engineering of eukaryotic microalgae[J]. Synthetic Biology Journal, 2023, 4(6): 1140-1160.

孙翰, 刘进. 真核微藻脂质代谢工程的研究进展和展望[J]. 合成生物学, 2023, 4(6): 1140-1160.

Figures/Tables 5

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门（分类）	种	基因库			参考文献
门（分类）	种	细胞核	线粒体	叶绿体	参考文献
绿藻门	Chromochloris zofingiensis SAG UTEX32	SRR5310949-5310954	SRR5310949-5310954	SRR5310949-5310954	[13]
	Ostreococcus tauri RCC1115	NERT00000000	—	—	[14]
	Botryococcus braunii	MVGU00000000	KR057902.1	KM462884.1	[15]
	Chlamydomonas reinhardtii CC503 cw92 mt+	ABCN00000000	U03843	BK000554.2	[16]
	Auxenochlorella pyrenoidosa FACHB-9	ANZC00000000	—	—	[17]
	Chlorella vulgaris	LDKB00000000	—	AB001684.1	[18]
	Floydiella terrestris	ADIC01000000	—	—	[19]
	Ostreococcus lucimarinus CCE9901	GCA_000092065.1	—	—	[20]
	Micromonas pusilla CCMP1545	ACCP00000000	FJ858268.1	FJ858269	[21]
	Micromonas sp. RCC299	GCA_000090985.2	FJ859351.1	FJ858267.1	[21]
	Volvox carteri Eve	ACJH00000000	—	—	[22]
	Ostreococcus tauri RCC 4221	CAID00000000	CR954200.2	CR954199.2	[23]
	Auxenochlorella protothecoides UTEX 2341/0710	MUYL00000000 APJO00000000	—	/KC63163 4.1	[24]
硅藻门	Phaeodactylum tricornutum CCAP1055/1	ABQD00000000	HQ840789.1	EF067920.1	[25]
硅藻门	Thalassiosira pseudonana CCMP1335	AAFD00000000	DQ186202.1	EF067921.1	[26]
红藻门	Cyanidioschzon merolae 10D	AP006483	D89861.1	AB002583.1	[27-28]
红藻门	Porphyridium purpureum NIES 2140	AROW00000000	—	AP012987.1	[29]
真眼点藻门	Nannochloropsis oceanica IMET1	MPCS00000000.1	KC598090.1	KC568462.1	[30]
真眼点藻门	Nannochloropsis gaditana CCMP526	AGNI00000000	KC012945.1	KC012944.1	[31]

门（分类）	种	基因库			参考文献
门（分类）	种	细胞核	线粒体	叶绿体	参考文献
绿藻门	Chromochloris zofingiensis SAG UTEX32	SRR5310949-5310954	SRR5310949-5310954	SRR5310949-5310954	[13]
	Ostreococcus tauri RCC1115	NERT00000000	—	—	[14]
	Botryococcus braunii	MVGU00000000	KR057902.1	KM462884.1	[15]
	Chlamydomonas reinhardtii CC503 cw92 mt+	ABCN00000000	U03843	BK000554.2	[16]
	Auxenochlorella pyrenoidosa FACHB-9	ANZC00000000	—	—	[17]
	Chlorella vulgaris	LDKB00000000	—	AB001684.1	[18]
	Floydiella terrestris	ADIC01000000	—	—	[19]
	Ostreococcus lucimarinus CCE9901	GCA_000092065.1	—	—	[20]
	Micromonas pusilla CCMP1545	ACCP00000000	FJ858268.1	FJ858269	[21]
	Micromonas sp. RCC299	GCA_000090985.2	FJ859351.1	FJ858267.1	[21]
	Volvox carteri Eve	ACJH00000000	—	—	[22]
	Ostreococcus tauri RCC 4221	CAID00000000	CR954200.2	CR954199.2	[23]
	Auxenochlorella protothecoides UTEX 2341/0710	MUYL00000000 APJO00000000	—	/KC63163 4.1	[24]
硅藻门	Phaeodactylum tricornutum CCAP1055/1	ABQD00000000	HQ840789.1	EF067920.1	[25]
硅藻门	Thalassiosira pseudonana CCMP1335	AAFD00000000	DQ186202.1	EF067921.1	[26]
红藻门	Cyanidioschzon merolae 10D	AP006483	D89861.1	AB002583.1	[27-28]
红藻门	Porphyridium purpureum NIES 2140	AROW00000000	—	AP012987.1	[29]
真眼点藻门	Nannochloropsis oceanica IMET1	MPCS00000000.1	KC598090.1	KC568462.1	[30]
真眼点藻门	Nannochloropsis gaditana CCMP526	AGNI00000000	KC012945.1	KC012944.1	[31]

藻种	株	细胞器	转化方法	启动子	报告基因（标记）	参考文献
Chlamydomonas reinhardtii	dw15.1	细胞核	电穿孔	Hsp70A-RBCS2	ble	[91]
	137c	原生质体	农杆菌转化	ChlL	nptⅡ (GUS)	[92]
Chlorella vulgaris	UMT-M1	细胞核	农杆菌转化	CaMV35S/OlexA-TATA	hpt (GFP)	[93]
	CBS 15-2075	叶绿体	PEG介导	CaMV35S	aphⅧ (EGFP)	[94]
	211/11B	细胞核	基因枪	CaMV35S	nptⅡ (GUS)	[95]
Chlorella ellipsoidea	SD0801	细胞核	PEG介导	Ubi-1 (Polyubiquitin-1)	ble (GUS)	[96]
Chlorella ellipsoidea	Nrm-4	细胞核	电穿孔	Ubi	nptⅡ	[97]
Chlorella sp.	DT	细胞核	PEG介导	Actin1/CaMV35S	hpt (GUS)	[98]
Chromochloris zofingiensis	ATCC 30412	细胞核	电穿孔/基因枪	PDS/NIT/RBCS2	pds	[99]
Nannochloropsis sp.	PP983	细胞核	电穿孔	TUB	ble (GUS)	[100]
	W2J3B	细胞核	电穿孔	VCP1/2	ble	[101]
Nannochloropsis salina	CCMP1776	细胞核	基因枪	TUB	ble	[102]
Nannochloropsis oceanica	IMET1	细胞核	电穿孔	β-tubulin	ble (GUS)	[103]
Nannochloropsis oculata	NIES-2146	细胞核	纤维素酶预处理和电穿孔	Hsp70A-RBCS2	CP (purple chromoprotein)	[104]
Nannochloropsis gaditana	CCMP526	细胞核	电穿孔	TUB/UEP/Hsp70A	ble	[31]
Dunaliella salina	—	细胞核	农杆菌转化	RBCS2/CaMV35S	hptⅡ (GFP)	[105]
	UTEX-1644	细胞核	玻璃珠法	Ubi-1	pat	[106]
	L1644	细胞核	基因枪	Actin	bar	[107]
Dunaliella tertiolecta	—	细胞核	玻璃珠法	CaMV35S	bar (EGFP)	[108]
	SE0045	叶绿体	基因枪	PsbD	ereB	[109]
Phaeodactylum tricornutum	NRIA-0065	细胞核	基因枪	DIV/FCP	ble (EGFP)	[110]
	CCMP2561	细胞核	电穿孔	FCPB	ble (GFP/GUS)	[111]
Thalassiosira pseudonana	CCMP1335	细胞核	基因枪	FCP2	ble (EGFP)	[112]
Haematococcus pluvialis	1844	细胞核	基因枪	PsaD	ble/pds	[113]
	CCAP 34/7	叶绿体	PEG介导	RBCL	aadA	[114]
	NIES-144	细胞核	基因枪	PDS	pds	[115]
	—	细胞核	农杆菌转化	CaMV35S	hpt (GFP)	[116]
Fistulifera solaris	JPCC DA0580	细胞核	基因枪	GAPDH	AphⅧ (G418) (GFP)	[62]
	JPCC DA0580	叶绿体	基因枪	CaMV35S/FCPB/RSV	nptⅡ (GFP)	[117]
Monoraphidium neglectum	SAG 48.87	细胞核	电穿孔	CAB2 (Chlorophyll a/b-binding protein)	AphⅧ (HygromycinB)	[118]
Neochloris oleoabundans	UTEX 1185	细胞核	电穿孔	β2-tubulin	Hyg3 (HygromycinB)	[119]
Botryococcus braunii	UTEX572	细胞核	纤维素酶预处理和电穿孔	CaMV35S	aphⅧ	[120]
Tetraselmis chuii	CCAP 66/21B	细胞核	农杆菌转化	CaMV35S	ble	[121]
Symbiodinium spp.	Mf11.5b.1	细胞核	玻璃珠法	CaMV35S	nptⅡ	[122]
Lobosphaera incisa	SAG 2468	细胞核	电穿孔	RBCS	ble	[123]
Isochrysis sp.	H-13	细胞核	农杆菌转化	LAT	pds	[124]
Parachlorella kessleri	—	细胞核	农杆菌转化	CaMV35S	hpt	[125]
Fistulifera sp.	JPCC DA0580	细胞核	基因枪	FCPB	nptⅡ	[117]
Aurantiochytrium sp.	KRS101	细胞核	电穿孔	GAP	CYH	[126]
Scenedesmus obliquus	FSP-3	细胞核	电穿孔	CaMV35S	CAT	[127]
Schizochytrium sp.	TIO1101	细胞核	农杆菌转化	TEF1/CaMV35S	nptⅡ	[128]
Platymonas subcordiformis	—	细胞核	玻璃珠法	CaMV35S	—	[129]
Porphyridium sp.	—	叶绿体	基因枪	AHAS	AHAS	[130]
Arthrospira sp.	PCC9438	细胞核	电穿孔	CAT	CAT	[131]

藻种	株	细胞器	转化方法	启动子	报告基因（标记）	参考文献
Chlamydomonas reinhardtii	dw15.1	细胞核	电穿孔	Hsp70A-RBCS2	ble	[91]
	137c	原生质体	农杆菌转化	ChlL	nptⅡ (GUS)	[92]
Chlorella vulgaris	UMT-M1	细胞核	农杆菌转化	CaMV35S/OlexA-TATA	hpt (GFP)	[93]
	CBS 15-2075	叶绿体	PEG介导	CaMV35S	aphⅧ (EGFP)	[94]
	211/11B	细胞核	基因枪	CaMV35S	nptⅡ (GUS)	[95]
Chlorella ellipsoidea	SD0801	细胞核	PEG介导	Ubi-1 (Polyubiquitin-1)	ble (GUS)	[96]
Chlorella ellipsoidea	Nrm-4	细胞核	电穿孔	Ubi	nptⅡ	[97]
Chlorella sp.	DT	细胞核	PEG介导	Actin1/CaMV35S	hpt (GUS)	[98]
Chromochloris zofingiensis	ATCC 30412	细胞核	电穿孔/基因枪	PDS/NIT/RBCS2	pds	[99]
Nannochloropsis sp.	PP983	细胞核	电穿孔	TUB	ble (GUS)	[100]
	W2J3B	细胞核	电穿孔	VCP1/2	ble	[101]
Nannochloropsis salina	CCMP1776	细胞核	基因枪	TUB	ble	[102]
Nannochloropsis oceanica	IMET1	细胞核	电穿孔	β-tubulin	ble (GUS)	[103]
Nannochloropsis oculata	NIES-2146	细胞核	纤维素酶预处理和电穿孔	Hsp70A-RBCS2	CP (purple chromoprotein)	[104]
Nannochloropsis gaditana	CCMP526	细胞核	电穿孔	TUB/UEP/Hsp70A	ble	[31]
Dunaliella salina	—	细胞核	农杆菌转化	RBCS2/CaMV35S	hptⅡ (GFP)	[105]
	UTEX-1644	细胞核	玻璃珠法	Ubi-1	pat	[106]
	L1644	细胞核	基因枪	Actin	bar	[107]
Dunaliella tertiolecta	—	细胞核	玻璃珠法	CaMV35S	bar (EGFP)	[108]
	SE0045	叶绿体	基因枪	PsbD	ereB	[109]
Phaeodactylum tricornutum	NRIA-0065	细胞核	基因枪	DIV/FCP	ble (EGFP)	[110]
	CCMP2561	细胞核	电穿孔	FCPB	ble (GFP/GUS)	[111]
Thalassiosira pseudonana	CCMP1335	细胞核	基因枪	FCP2	ble (EGFP)	[112]
Haematococcus pluvialis	1844	细胞核	基因枪	PsaD	ble/pds	[113]
	CCAP 34/7	叶绿体	PEG介导	RBCL	aadA	[114]
	NIES-144	细胞核	基因枪	PDS	pds	[115]
	—	细胞核	农杆菌转化	CaMV35S	hpt (GFP)	[116]
Fistulifera solaris	JPCC DA0580	细胞核	基因枪	GAPDH	AphⅧ (G418) (GFP)	[62]
	JPCC DA0580	叶绿体	基因枪	CaMV35S/FCPB/RSV	nptⅡ (GFP)	[117]
Monoraphidium neglectum	SAG 48.87	细胞核	电穿孔	CAB2 (Chlorophyll a/b-binding protein)	AphⅧ (HygromycinB)	[118]
Neochloris oleoabundans	UTEX 1185	细胞核	电穿孔	β2-tubulin	Hyg3 (HygromycinB)	[119]
Botryococcus braunii	UTEX572	细胞核	纤维素酶预处理和电穿孔	CaMV35S	aphⅧ	[120]
Tetraselmis chuii	CCAP 66/21B	细胞核	农杆菌转化	CaMV35S	ble	[121]
Symbiodinium spp.	Mf11.5b.1	细胞核	玻璃珠法	CaMV35S	nptⅡ	[122]
Lobosphaera incisa	SAG 2468	细胞核	电穿孔	RBCS	ble	[123]
Isochrysis sp.	H-13	细胞核	农杆菌转化	LAT	pds	[124]
Parachlorella kessleri	—	细胞核	农杆菌转化	CaMV35S	hpt	[125]
Fistulifera sp.	JPCC DA0580	细胞核	基因枪	FCPB	nptⅡ	[117]
Aurantiochytrium sp.	KRS101	细胞核	电穿孔	GAP	CYH	[126]
Scenedesmus obliquus	FSP-3	细胞核	电穿孔	CaMV35S	CAT	[127]
Schizochytrium sp.	TIO1101	细胞核	农杆菌转化	TEF1/CaMV35S	nptⅡ	[128]
Platymonas subcordiformis	—	细胞核	玻璃珠法	CaMV35S	—	[129]
Porphyridium sp.	—	叶绿体	基因枪	AHAS	AHAS	[130]
Arthrospira sp.	PCC9438	细胞核	电穿孔	CAT	CAT	[131]

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