Molecular characterization of flavanone 3-hydroxylase gene and flavonoid accumulation in two chemotyped safflower lines in response to methyl jasmonate stimulation.
Among secondary metabolites, flavonoids are particularly crucial for plant growth, development, and reproduction, as well as beneficial for maintenance of human health. As a flowering plant, safflower has synthesized a striking variety of flavonoids with various pharmacologic properties. However, far less research has been carried out on the genes involved in the biosynthetic pathways that generate these amazing flavonoids, especially characterized quinochalcones. In this study, we first cloned and investigated the participation of a presumed flavanone 3-hydroxylase gene (F3H) from safflower (CtF3H) in a flavonoid biosynthetic pathway.



Bioinformation analysis showed that CtF3H shared high conserved residues and confidence with F3H from other plants. Subcellular localization uncovered the nuclear and cytosol localization of CtF3H in onion epidermal cells. The functional expressions of CtF3H in Escherichia coli BL21(DE3)pLysS cells in the pMAL-C5x vector led to the production of dihydrokaempferol when naringenin was the substrate. Furthermore, the transcriptome expression of CtF3H showed a diametrically opposed expression pattern in a quinochalcone-type safflower line (with orange-yellow flowers) and a flavonol-type safflower line (with white flowers) under external stimulation by methyl jasmonate (MeJA), which has been identified as an elicitor of flavonoid metabolites. Further metabolite analysis showed the increasing tendency of quinochalcones and flavonols, such as hydroxysafflor yellow A, kaempferol-3-O-β-D-glucoside, kaempferol-3-O-β-rutinoside, rutin, carthamin, and luteolin, in the quinochalcone-type safflower line. Also, the accumulation of kaempferol-3-O-β-rutinoside and kaempferol-3-O-β-D-glucoside in flavonols-typed safflower line showed enhanced accumulation pattern after MeJA treatment. However, other flavonols, such as kaempferol, dihydrokaempferol and quercetin-3-O-β-D-glucoside, in flavonols-typed safflower line presented down accumulation respond to MeJA stimulus.



Our results showed that the high expression of CtF3H in quinochalcone-type safflower line was associated with the accumulation of both quinochalcones and flavonols, whereas its low expression did not affect the increased accumulation of glycosylated derivatives (kaempferol-3-O-β-rutinoside and rutin) in flavonols-typed safflower line but affect the upstream precursors (D-phenylalanine, dihydrokaempferol, kaempferol), which partly revealed the function of CtF3H in different phenotypes and chemotypes of safflower lines.
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China.
YanHua
Y
Tu
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China.
Fei
F
Liu
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China.
DanDan
D
Guo
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China.
LiJiao
L
Fan
School of Biological and Environmental Sciences, Nanjing Forestry University, Nanjing, 210095, People's Republic of China.
ZhenXian
Z
Zhu
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China.
YingRu
Y
Xue
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China. gaoyue2000@hotmail.com.
Yue
Y
Gao
School of Pharmacy, Second Military Medical University, Shanghai, 200433, People's Republic of China. mlguo@126.com.
MeiLi
M
Guo
D000085:Acetates / Q000494:pharmacology*; D000595:Amino Acid Sequence; D031187:Carthamus tinctorius / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology*; D005419:Flavonoids / Q000096:biosynthesis*; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D054883:Oxylipins / Q000494:pharmacology*; D010937:Plant Growth Regulators / Q000494:pharmacology*; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment
D000085:Acetates; D003517:Cyclopentanes; D005419:Flavonoids; D054883:Oxylipins; D010937:Plant Growth Regulators; D010940:Plant Proteins; C072239:methyl jasmonate; D006899:Mixed Function Oxygenases; C048797:flavanone 3-dioxygenase
Flavanone 3-hydroxylase gene; Flavonoid accumulation; Functional characterization; Methyl jasmonate; Safflower (Carthamus tinctorius L.)