title: RNAi-mediated suppression of dihydroflavonol 4-reductase in tobacco allows fine-tuning of flower color and flux through the flavonoid biosynthetic pathway.
abstract: To examine flux regulation in the flavonoid pathway of tobacco flowers, we suppressed two genes for dihydroflavonol 4-reductase (NtDFR 1 and 2) by RNA interference (Ri)-mediated post transcriptional gene silencing in pink-flowered tobacco. Two phenotypes were observed, pale pink (DFR-Ri_PP)- and white (DFR-Ri_W)-flowered lines. The relative mRNA levels of NtDFR genes in DFR-Ri_PP and DFR-Ri_W lines were reduced by 79%-95% relative to non-transformed (NT) plants. DFR-Ri_W lines had five-fold higher levels of small interference RNAs compared to DFR-Ri_PP lines. Expression of eight structural genes in the flavonoid pathway was significantly increased in DFR-Ri_W lines but not in DFR-Ri_PP lines based on quantitative RT-PCR. Anthocyanin contents correlated with flower color, with a reduction of 72%-97% in DFR-Ri_PP and DFR-Ri_W lines. Decreases in anthocyanin in flower were proportional with reductions of proanthocyanidin content in seeds. Two pale pink lines, DFR-Ri_PP 17 and 20, with anthocyanin decreases and the lowest level of DFR gene silencing, had higher (dihydro) flavonol production than a white flowered line, DFR-Ri_W 67. This finding suggests that suppression of DFR can increase the total levels of flavonoids due to (dihydro) flavonol biosynthesis. Our observations that higher suppression of DFR had a greater influence on the expression of flavonoid biosynthetic genes demonstrates the key role of DFR in the pathway and allows selection among DFR-Ri lines for plants with specific gene expression profiles to fine-tune flux through the pathway.
authors.affiliation: National Academy of Agricultural Science, Rural Development Administration, Jeonju, 54874, Republic of Korea. Electronic address: limsh2@korea.kr.
authors.forename: Sun-Hyung
authors.initials: SH
authors.lastname: Lim
authors.affiliation: Department of Genetic Engineering and Graduate School of Biotechnology, Kyung Hee University Graduate School of Biotechnology, Kyung Hee University, Yongin, 17104, Republic of Korea.
authors.forename: Min-Kyung
authors.initials: MK
authors.lastname: You
authors.affiliation: National Academy of Agricultural Science, Rural Development Administration, Jeonju, 54874, Republic of Korea.
authors.forename: Da-Hye
authors.initials: DH
authors.lastname: Kim
authors.affiliation: Division of Life Sciences and Bio-Resource and Environmental Center, Incheon National University, Incheon, 22012, Republic of Korea.
authors.forename: Jae Kwang
authors.initials: JK
authors.lastname: Kim
authors.affiliation: National Academy of Agricultural Science, Rural Development Administration, Jeonju, 54874, Republic of Korea.
authors.forename: Jong-Yeol
authors.initials: JY
authors.lastname: Lee
authors.affiliation: Department of Genetic Engineering and Graduate School of Biotechnology, Kyung Hee University Graduate School of Biotechnology, Kyung Hee University, Yongin, 17104, Republic of Korea. Electronic address: sunhwa@khu.ac.kr.
authors.forename: Sun-Hwa
authors.initials: SH
authors.lastname: Ha
mesh_terms: D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors* / Q000235:genetics*; D053898:Biosynthetic Pathways / Q000235:genetics; D005419:Flavonoids / Q000096:biosynthesis*; D035264:Flowers / Q000235:genetics / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D010858:Pigmentation / Q000235:genetics; D010940:Plant Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics*; D030821:Plants, Genetically Modified; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism*
chemical_list: D005419:Flavonoids; D010940:Plant Proteins; D012333:RNA, Messenger; D018749:RNA, Plant; D000429:Alcohol Oxidoreductases; C056570:dihydroflavanol 4-reductase
keywords: Anthocyanin; DFR; FLS; Metabolic flux; Post transcriptional gene silencing; Proanthocyanidin; Tobacco