Expression Profiling of Regulatory and Biosynthetic Genes in Contrastingly Anthocyanin Rich Strawberry ( <i>Fragaria</i> × <i>ananassa</i> ) Cultivars Reveals Key Genetic Determinants of Fruit Color

Hossain, Mohammad Rashed,Kim, Hoy-Taek,Shanmugam, Ashokraj,Nath, Ujjal Kumar,Goswami, Gayatri,Song, Jae-Young,Park, Jong-In,Nou, Ill-Sup MDPI 2018 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.19 No.3

<P>Anthocyanins are the resultant end-point metabolites of phenylapropanoid/flavonoid (F/P) pathway which is regulated at transcriptional level via a series of structural genes. Identifying the key genes and their potential interactions can provide us with the clue for novel points of intervention for improvement of the trait in strawberry. We profiled the expressions of putative regulatory and biosynthetic genes of cultivated strawberry in three developmental and characteristically colored stages of fruits of contrastingly anthocyanin rich cultivars: Tokun, Maehyang and Soelhyang. Besides <I>FaMYB10,</I> a well-characterized positive regulator, <I>FaMYB5</I>, <I>FabHLH3</I> and <I>FabHLH3-delta</I> might also act as potential positive regulators, while <I>FaMYB11</I>, <I>FaMYB9</I>, <I>FabHLH33</I> and <I>FaWD44-1</I> as potential negative regulators of anthocyanin biosynthesis in these high-anthocyanin cultivars. Among the early BGs, <I>Fa4CL7</I>, <I>FaF3H</I>, <I>FaCHI1</I>, <I>FaCHI3</I>, and <I>FaCHS,</I> and among the late BGs, <I>FaDFR4-3</I>, <I>FaLDOX</I>, and <I>FaUFGT2</I> showed significantly higher expression in ripe fruits of high anthocyanin cultivars Maehyang and Soelhyang. Multivariate analysis revealed the association of these genes with total anthocyanins. Increasingly higher expressions of the key genes along the pathway indicates the progressive intensification of pathway flux leading to final higher accumulation of anthocyanins. Identification of these key genetic determinants of anthocyanin regulation and biosynthesis in Korean cultivars will be helpful in designing crop improvement programs.</P>

매향 딸기로부터 anthocyanin 합성 유전자의 분리 및 과실발달 과정에서의 발현 분석

배기석(Kisuk Bae),길준영(Joon Yeong Kihl),피재호(Jaeho Pyee) 한국생명과학회 2008 생명과학회지 Vol.18 No.2

‘매향’ 딸기의 안토시아닌 생합성은 개화 후 26일째 시작되어 과실의 성숙기 동안 계속된다. 딸기로부터 안토시아닌의 생합성에 관여하는 주요 유전자를 분리하였다. 각각의 유전자에 대해, 다양한 식물체의 유사 유전자의 염기서열을 비교하여 PCR (polymerase chain reaciton) primer를 제작하였다. 숙기의 딸기에서 분리된 total RNA로부터 합성된 cDNA와각 primer를 이용하여 RT (reverse transcriptase)-PCR을 수행하였다. 각 cDNA clone의 염기서열을 작성하여 분석한 결과, 이들은 안토시아닌 생합성에 관여하는 phenylalanine ammonialyase (PAL), 4-cummarate CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone-3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidine synthase (ANS) 그리고 UDP-glucose: flavonoid-3-O-glucosyltransferase (UFGT) 효소에 해당되었다. Northern blot 분석 결과, 이들 유전자는 과실 발달 과정에서 시기적으로 조절되었다. 특히 PAL을 제외한 모든 유전자는 과실에서만 주로 발현되었다. PAL, DFR 그리고 ANS 유전자는 과실 초기 발달 단계인 개화 후 10일에 검출된 후 감소하다가, 22일에 다시 증가하기 시작하여 34일에 최대가 되었다. 한편, 다른 유전자들은 초기에는 발현되지 않다가, 안토시아닌이 축적되기 시작하는 개화 후 22~30일에 처음으로 검출되었다. 본 연구를 통해, 딸기 과실 발달과정에서 안토시아닌 생합성 과정에 관여하는 여러 유전자가 과실 숙기에 함께 조절되는 현상을 알 수 있다. 이러한 연구 결과는 안토시아닌 합성과정을 제어하는 조절 유전자가 존재한다는 것을 시사한다. 그리고 딸기의 안토시아닌 생합성 유전자의 발현 패턴을 크게 두 가지로 나눌 수 있는 것으로 보아, 딸기의 안토시아닌 생합성에는 적어도 두 가지 서로 다른 조절 기작이 관여하여 색소 발달 과정을 제어할 것으로 보인다. Anthocyanin synthesis in strawberry (Fragaria x ananassa cv Maehyang) begins approximately 26 days postflowering and continued throughout fruit ripening. A set of cDNA clones encoding the anthocyanin biosynthetic enzymes were isolated from strawberry. A pair of primers were designed for polymerase chain reaction (PCR) through the comparison of the nucleotide sequences of homologous genes from diverse plants. Reverse transcriptase-PCRs were performed using cDNA synthesized from ripe fruit total RNA and the primers corresponding to each gene. Eight genes of the anthocyanin pathway were cloned and confirmed by sequencing to code for phenylalanine ammonia lyase (PAL), 4-cummarate CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone-3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidine synthase (ANS), UDP-glucose:flavonoid-3-O-glucosyltransferase (UFGT). Northern analyses showed that the corresponding genes were differentially ex-pressed during the fruit development process. All genes except PAL were predominantly expressed in fruit. Expression of PAL, DFR and ANS was detected 10 days postflowering at the early stage of fruit development, declined for a while and sharply increased 22 days postflowering then showed a peak 34 days postflowering. The other genes, however, were not expressed up to 22 or 30 days post-flowering when the initial fruit ripening events occur at the time of initiation of anthocyanin accumulation. The onset of anthocyanin synthesis in ripening strawberry coincides with a coordinated induction of the anthocyanin pathway genes, suggesting the involvement of regulatory genes. We pro-pose that at least two different regulatory mechanisms play a role in the biosynthesis of anthocyanin during color development of strawberry.

Expression of anthocyanin biosynthesis-related genes reflects the peel color in purple tomato

강송이,Md Abdur Rahim,Khandker Shazia Afrin,정희정,김효택,박종인,노일섭 한국원예학회 2018 Horticulture, Environment, and Biotechnology Vol.59 No.3

Tomato (Solanum lycopersicum L.) is one of the most important and popular vegetables worldwide. A wide range of tomato cultivars with different colored fruits is presently available. The purple peel of tomato fruit is due to the accumulation anthocyanin pigments, which are beneficial to both plants and humans. Cultivated tomatoes normally do not make anthocyanin in their fruit peel, but there are some wild relatives of cultivated tomato like Solanum chilense, S. habrochaites, S. cheesmanii, and S. lycopersicoides that do produce anthocyanins in their fruits. In this study, the purple fruit color was obtained by crossing ‘OSU blue’ (blue fruit) and ‘Purple mini’ (brown fruit) and subsequent self-pollination. Anthocyanins are produced via the flavonoid pathway and are regulated by the transcriptional complex of MYB, bHLH, and WD40 repeats transcription factors. We determined the expression profiles of genes related to anthocyanin biosynthesis in tomato genotypes with distinct fruit colors by qRT-PCR. Both the early and late biosynthetic genes of the anthocyanin pathway were up-regulated in the peels of purple tomato fruits, except Sl5GT. Moreover, the expression of the regulatory genes SlANT1 and SlAN1 was dramatically increased in the peels of purple tomato fruits. These results indicate that SlANT1 and SlAN1 might play an important role on anthocyanin biosynthesis in the peels of purple-fruited tomatoes via up-regulation of structural genes in the anthocyanin pathway.

Abscisic acid(ABA) 및 fluridone의 처리가 ‘Hongro’ 사과의 과피 착색에 미치는 영향

류수현(Suhyun Ryu),권용희(Yong Hee Kwon),도경란(Gyeong Ran Do),정재훈(Jae Hoon Jeong),한현희(Hyun Hee Han),한점화(Jeom Hwa Han) (사)한국생물환경조절학회 2016 생물환경조절학회지 Vol.25 No.4

식물 호르몬의 일종인 ABA의 처리가 사과 과피의 착색에 미치는 영향을 알아보기 위하여, 착색이 시작되는 시기의 ‘Hongro’ 사과에 abscisic acid (ABA)와 ABA의 생합성 저해제인 fluridone (FD)을 처리한 뒤 과피의 색변화를 관찰하였다. 실험 결과, ABA와 FD 처리에 의해 ‘Hongro’ 사과의 착색이 큰 영향을 받지 않았다. 과피의 붉은 색을 나타내는 hunter a값이 처리 7일 후까지 처리간에 차이가 없었다. 실제 과피의 안토시아닌 함량은 과피의 hunter a값의 변화와 같은 경향을 보이며 증가하였으며, ABA나 FD의 처리가 과피의 안토시아닌 축적에 영향을 주지 않았다. ABA의 처리 직후 과피의 내생 ABA함량이 급격히 증가하였고 처리 종료 시까지 높게 유지되었다. FD 처리구의 과피 내 ABA함량은 처리 6시간 후부터 대조구보다 낮아지다가, 처리 4일 후까지 지속적으로 감소하였다. ABA의 처리에 의해서 과피의 MdNCED2 (ABA 생합성 유전자)의 발현량이 증가하였고, MdACO1 (ethlyene 생합성 유전자) 및 안토시아닌 생합성 유전자 중 MdCHS와 MdDFR의 발현량이 증가하였다. 하지만 MdUFGT의 발현량은 ABA 처리에 의해서 변화가 없었다. The objective of this study was to determine the effect of exogenous abscisic acid (ABA) on the red coloration and endogenous ABA contents of apple fruit skins. ABA and fluridone (an ABA synthetic inhibitor, FD) was sprayed on ‘Hongro’ apple fruit skins at 107 days after full bloom (DAFB). Visual coloration and hunter’s color values were not affected by the ABA and FD treatments. Anthocyanin contents in fruit skins increased similarly to hunter a<SUP>*</SUP> values of fruit skins, but ABA and FD did not affect its accumulations. Liquid chromatography analysis revealed that endogenous ABA contents in control fruit increased at first and then decreased from 12 hours after the treatment. ABA treatment increased ABA contents in fruit skins from 2 hour after the treatment and it lasted until the end of the treatments. FD decreased ABA contents in fruit skins from 6 hours after the treatment. ABA treatment increased MdNCED2 (an ABA biosynthetic gene), MdACO1 (an ethylene biosynthetic gene), and MdCHS and MdDFR expressions. However, MdUFGT expressions were not affected by ABA treatment.

야간 고온에 의한 사과 후지 품종의 생리반응 및 과실품질 변화

류수현(Suhyun Ryu),권용희(YongHee Kwon),도경란(Kyeong Ran Do),한점화(Jeom Hwa Han),한현희(Hyun Hee Han),이한찬(Han Chan Lee) (사)한국생물환경조절학회 2015 생물환경조절학회지 Vol.24 No.3

최근 지구 온난화로 인해 열대야 발생 빈도가 증가하고 있으며, 야간 저온의 감소에 의한 사과의 착색 불량이 예상된다. 따라서 본 연구는 생육기 후반의 야간 고온이 ‘후지’ 사과에 미치는 영향을 구명하기 위해 성숙기 과실 품질과 시기별 안토시아닌 생합성 유전자의 발현량을 조사하였고, 잎과 과실 내 당 함량을 비교하여 야간 고온에 의한 생리반응을 구명하고자 실시하였다. 야간온도는 7월부터 10월까지 인공기상실 내부에서 처리하였고, 대기온도 대비 -4℃, +4℃ 로 각각 설정하였다. 야간 고온은 과실의 횡경, 과중, 당도에 영향을 미치지 않았고 과피의 착색을 불량하게 하였지만, 착색시기의 안토시아닌 생합성 유전자의 발현량을 변화시키지 않았다. 또한, 야간 고온에 의해서 잎의 sorbitol, glucose 함량과 과실의 sorbitol, sucrose 함량이 대조구에 비해 감소하였고, 잎 조직을 구성하고 있는 책상, 해면 등의 광합성 조직에는 차이가 나타나지 않았지만 광합성 조직 내의 전분 함량이 감소하였다. 따라서 야간고온은 ‘후지’ 사과의 과실 품질에는 영향을 미치지 않았지만, 잎과 과실의 당 조성을 변화시켰으며, 안토시아닌 생합성 유전자의 발현과는 관계없이 과피의 착색을 억제하였다. Tropical night phenomenon has been increasing due to global warming recently, it is expected that fruit quality of apples will decrease due to elevated night temperature condition. In the present study, fruit quality at maturity, periodic anthocyanin biosynthetic gene expression and sugar contents in leaves and fruit flesh were investigated to establish the physiological responses of ‘Fuji’ apples under high night temperature. The night temperature were treated with such as ambient (control), ambient -4℃, and ambient +4℃. After the treatment, high night temperature didn’t affect fruit diameter, weight, and soluble sugar contents. Coloration of ambient +4℃ was poor than that of control, however there was no significant difference between these genes expression of control and that of ambient +4℃ treatment in the late coloration season. Increase of sorbitol and glucose contents at ambient +4℃ in leaves were smaller than those at control, and then sorbitol and sucrose contents in fruit flesh at ambient +4℃ were smaller than those at control. The cross section of leaves showed that there were no differences with the structure of parenchyma and epidermis tissues between the treatments, but starch granules in the palisade parenchyma cells decreased in high night temperature treatments. Consequently, high night temperature didn’t affect the fruit quality, but changed sugar contents in leaves and fruit flesh, and suppressed coloration regardless of anthocyanin biosynthetic gene expression.