Fruit Anthocyanin Research Articles

Molecular characterization of anthocyanin and betulinic acid biosynthesis in red and white mulberry fruits using high-throughput sequencing

To better understand the molecular mechanism of color formation in different varieties of the mulberry fruit, we investigated the functional genes related to anthocyanin and betulinic acid biosynthesis using high-throughput transcriptome sequencing and detected the primary and secondary metabolites in the white (Morus alba L. cv. ‘Turkey’) and red (Morus alba L. cv. ‘Cheongil’) mulberry cultivars. We obtained 171,702,058 high-quality reads with an average read length of 125 bp. These reads were assembled into 51,272 and 51,159 unigenes in Turkey and Cheongil, respectively. We also identified the genes related to anthocyanin and triterpene biosynthesis and investigated their expression and metabolite profiles. Overall, our transcriptome sequencing provides valuable information that could be used in gene discovery, marker-assisted selection, and investigation of metabolic pathways in mulberry. Additionally, gene expression and metabolite profiles provide new insights into the underlying mechanism of anthocyanin and betulinic acid biosynthesis and relationship between primary and secondary metabolites.

DFR and PAL gene transcription and their correlation with anthocyanin accumulation in Rhodomyrtus tomentosa (Aiton.) Hassk.

Abstract Objectives Rhodomyrtus tomentosa (Aiton.) Hassk. (R. tomentosa) is rich in nutrients and has multiple pharmacological applications. Anthocyanins confer color to the flowers and berries of R. tomentosa and provide protection against photodamage. The dihydroflavonol 4-reductase gene (DFR) and phenylalanine ammonialyase gene (PAL) are crucial for anthocyanin synthesis. Methods DFR and PAL transcript levels and anthocyanin content in the pigmented organs of R. tomentosa were investigated through qRT-PCR analysis and spectrophotometry, respectively. The glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was selected as the reference gene for the normalization of DFR and PAL transcript levels. Results Transcript levels of DFR and PAL were higher in organs with vigorous metabolism than those in senescent organs. DFR and PAL transcript levels were up-regulated during the initial and middle-maturity periods of fruit. These expression patterns are consistent with fruit color development. The highest transcript levels of PAL and DFR were observed during the middle-maturity period or the red-fruit period. Conclusion During the late maturity period of R. tomentosa fruit, the transcript levels of the two genes were down-regulated even though anthocyanins were continuously accumulated, which was different from the accumulation of anthocyanins in some late mature fruits.

Cyanidin-3-glucoside attenuates the angiogenesis of breast cancer via inhibiting STAT3/VEGF pathway.

Angiogenesis plays a pivotal role in breast cancer progression. Cyanidin-3-glucoside (C3G), one of the most widely distributed anthocyanins in edible fruits, shows antioxidative and anti-inflammatory property as well as induction of breast cancer cells apoptosis. However, the effect of C3G on breast cancer-induced angiogenesis remains unknown. In the present study, we found that C3G could attenuate breast cancer-induced angiogenesis via inhibiting VEGF, a key cytokine for angiogenesis, expression and secretion. Furthermore, signal transducer and activator of transcription 3 (STAT3) could transcriptionally activate VEGF, and C3G reduced STAT3 expression at both mRNA and protein level. Subsequently, our data showed that C3G induced miR-124 expression. Moreover, miR-124 could directly repress STAT3 expression, and miR-124-mediated STAT3 down-regulation was responsible for the inhibition of C3G on VEGF and angiogenesis. Taken together, we supplied more evidence to the anti-breast cancer property of C3G.

Pomological and phytochemical diversity in Iranian populations of Caucasian whortleberry (Vaccinum arctostaphylos L.)

Caucasian whortleberry (Vaccinium arctostaphylos L.), locally named Qare-Qat, is a shrub native to humid Caucasus area including northwest of Iran. It is a medicinal plant which has been intensively utilized in Iranian folk medicine as an antihypertensive and antidiabetic agent for many years. In order to determine the pomological and phytochemical properties of various caucasian whortleberry populations, aerial parts samples were collected from 11 locations in native regions. Current study indicated significant differences in pomological and phytochemical characteristics of Caucasian whortleberry (V. arctostaphylos L.) populations collected from 11 locations in 3 different province. Our results showed that the highest berry and flesh weight, number of seeds per berry, berry length and width, number of cluster per inflorescence and flowering stem length were observed from plants collected from Saghezchi-A population in Ardabil province, while the lowest values were obtained from Zendaneh population in Gilan province. In addition, the highest content of anthocyanin, total phenolics and antioxidant in both fruits and leaves of V. arctostaphylos L. were mainly recorded from Saghezchi-A population followed by Khanghah population. According to studied traits, V. arctostaphylos L. populations were divided into three different groups. Saghezchi-A, Khanghah and Saghezchi-A populations were placed in a same group with mainly the best pomological and phytochemical properties. These locations were characterized by high solar radiation. Therefore, they can be exploited for selection of suitable genotypes for organizing the berry breeding programs and taking advantage of this plant in garden establishment and fruit production investigations.

Massive phenotyping of multiple cranberry populations reveals novel QTLs for fruit anthocyanin content and other important chemical traits.

Because of its known phytochemical activity and benefits for human health, American cranberry (Vaccinium macrocarpon L.) production and commercialization around the world has gained importance in recent years. Flavonoid compounds as well as the balance of sugars and acids are key quality characteristics of fresh and processed cranberry products. In this study, we identified novel QTL that influence total anthocyanin content (TAcy), titratable acidity (TA), proanthocyanidin content (PAC), Brix, and mean fruit weight (MFW) in cranberry fruits. Using repeated measurements over the fruit ripening period, different QTLs were identified at specific time points that coincide with known chemical changes during fruit development and maturation. Some genetic regions appear to be regulating more than one trait. In addition, we demonstrate the utility of digital imaging as a reliable, inexpensive and high-throughput strategy for the quantification of anthocyanin content in cranberry fruits. Using this imaging approach, we identified a set of QTLs across three different breeding populations which collocated with anthocyanin QTL identified using wet-lab approaches. We demonstrate the use of a high-throughput, reliable and highly accessible imaging strategy for predicting anthocyanin content based on cranberry fruit color, which could have a large impact for both industry and cranberry research.

The atroviolacea Gene Encodes an R3-MYB Protein Repressing Anthocyanin Synthesis in Tomato Plants.

The anthocyanin biosynthetic pathway is well characterized in plants. However, in tomato (Solanum lycopersicum L.) an exhaustive knowledge of its regulation is still lacking. Tomato mutants showing higher levels of anthocyanins in fruits or vegetative tissues, such as Anthocyanin fruit (Aft) or atroviolacea (atv), have been extensively exploited in the attempt to clarify the process. Nevertheless, only candidate genes have been proposed as responsible for such phenotypes. The recessive atv mutation likely represents an allelic variant of a gene introgressed in tomato from wild Solanum species. We performed genome sequencing of atv/atv plants followed by candidate gene analysis, and identified a mutated gene encoding an R3-MYB protein. When overexpressed, this protein abolished anthocyanin production in tomato seedlings and plants, by silencing key regulators and biosynthetic genes of the pathway. The functional analysis of the protein clearly showed that it can negatively interfere with the activation of the anthocyanin biosynthetic pathway mediated by the endogenous MYB-bHLH-WDR (MBW) complexes. In particular, this R3-MYB protein can directly bind the bHLH factors which are part of the MBW complexes, therefore acting as a competitive inhibitor. The R3-MYB protein here described is therefore involved in a feedback mechanism that dampens the production of anthocyanins once activated by endogenous or exogenous stimuli. The atv mutation causes the production of a truncated version of the R3-MYB factor that cannot retain the full potential to inhibit the MBW complexes, thus leading to a constitutively higher production of anthocyanins.

Transcriptome sequencing reveals role of light in promoting anthocyanin accumulation of strawberry fruit

The woodland strawberry, Fragaria vesca, is a model plant for the diverse Rosaceae family, which contains many valuable fruit and ornamental crops. Light is an important external environment factor which influences all aspects of plant growth and development, including fruit ripening. Transcription regulation of light will provide insights into effect of light for fruit ripening. We treat strawberry fruit with dark or light, and by the RNA-Seq method, compare transcriptional level between dark-treated and light-treated strawberry fruits. Additionally, we detect anthocyanin and sugar accumulation in two-treated conditions of fruits. Moreover, we detect the protein change of FvMYB10, a key regulator of anthocyanin biosynthesis, under dark and light. Light can promote anthocyanin and soluble sugar accumulation in mature fruit, and also increases the expression of aroma-related genes and stabilizes FvMYB10 protein. Our findings reveal that light is essential for anthocyanin and sugar accumulation and regulates FvMYB10 at transcriptional level and post-translation level.

Tomato SlAN11 regulates flavonoid biosynthesis and seed dormancy by interaction with bHLH proteins but not with MYB proteins

The flavonoid compounds are important secondary metabolites with versatile human nutritive benefits and fulfill a multitude of functions during plant growth and development. The abundance of different flavonoid compounds are finely tuned with species-specific pattern by a ternary MBW complex, which consists of a MYB, a bHLH, and a WD40 protein, but the essential role of SlAN11, which is a WD40 protein, is not fully understood in tomato until now. In this study, a tomato WD40 protein named as SlAN11 was characterized as an effective transcription regulator to promote plant anthocyanin and seed proanthocyanidin (PA) contents, with late flavonoid biosynthetic genes activated in 35S::SlAN11 transgenic lines, while the dihydroflavonol flow to the accumulation of flavonols or their glycosylated derivatives was reduced by repressing the expression of SlFLS in this SlAN11-overexpressed lines. The above changes were reversed in 35S::SlAN11-RNAi transgenic lines except remained levels of flavonol compounds and SlFLS expression. Interestingly, our data revealed that SlAN11 gene could affect seed dormancy by regulating the expressions of abscisic acid (ABA) signaling-related genes SlABI3 and SlABI5, and the sensitivity to ABA treatment in seed germination is conversely changed by SlAN11-overexpressed or -downregulated lines. Yeast two-hybrid assays demonstrated that SlAN11 interacted with bHLH but not with MYB proteins in the ternary MBW complex, whereas bHLH interacted with MYB in tomato. Our results indicated that low level of anthocyanins in tomato fruits, with low expression of bHLH (SlTT8) and MYB (SlANT1 and SlAN2) genes, remain unchanged upon modification of SlAN11 gene alone in the transgenic lines. These results suggest that the tomato WD40 protein SlAN11, coordinating with bHLH and MYB proteins, plays a crucial role in the fine adjustment of the flavonoid biosynthesis and seed dormancy in tomato.

Визначення вмісту антоціанів і танінів у аронії чорноплідної плодах

Introduction. Aronia melanocarpa is widely cultivated in Ukraine as a food, medicinal and ornamental plant. The quality of fresh fruit of Aronia melanocarpa was regulated by the requirements of the pharmaceutical article 42-66-87 "Fresh fruit of Aronia melanocarpa", but the article required a revision using modern approaches to the standardization of medicinal plant material.The aim of the study – determination of the content of anthocyanins and tannins in fresh and dried fruit of Aronia melanocarpa for inclusion the results of the research in the section of the monograph "Quantitative determination".Research Methods. The determination of the content of anthocyanins and tannins in fresh and dried fruit of Aronia melanocarpa was carried out by the method of absorption spectrophotometry.Results and Discussion. The content of anthocyanins in fresh fruit of Aronia melanocarpa is from (0.45±0.01) % to (0.56±0.02) % in terms of cyanidin-3-O-glucoside chloride, the content of tannins in dried fruit of Aronia melanocarpa is from (1.51±0.02) % to (2.41±0.03) % in terms of pyrogallol and dry raw materials. Therefore we proposed to enter the indicator: the content of anthocyanins – at least 0.40 % in terms of cyanidin-3-O-glucoside chloride to the national monograph "Fresh fruit of Aronia melanocarpaN", and the indicator: the content of tannins – not less than 1.5 % in terms of pyrogallol and dry raw materials to the national monograph "Dried fruit of Aronia melanocarpaN".Conclusion. By the method of absorption spectrophotometry in fresh fruit of Aronia melanocarpa the content of anthocyanins was determined, in dried fruit of Aronia melanocarpa – tannins. The results of the research were used during the development of the national monographs "Fresh fruit of Aronia melanocarpaN" and "Dried fruit of Aronia melanocarpaN".

Comparative transcriptome analysis of genes involved in anthocyanin synthesis in blueberry

Blueberry (Vaccinium, family Ericaceae) is well known for its strong antioxidant properties and abundant active ingredients including anthocyanins, flavonols, and proanthocyanidins. In this study, variations in anthocyanin and phenolic compounds content in Bluecrop and Northblue blueberry cultivar fruits were studied, and comparative transcriptome analysis was performed to analyze differences in the molecular mechanisms of anthocyanin biosynthesis. A total of 13 799 unique genes were identified by differential expression analysis, and further subjected to GO classification and pathway enrichment. Nine differentially expressed genes (DEGs), including CHI, DFR, F3′H, FLS, CHS, OMT, UGT, ANS and F3H, were selected to validate the differential expression data using quantitative real-time PCR. The obtained qRT-PCR expression results were consistent with the RNA-Seq results. The expression levels of 9 candidate genes involved in flavonoid biosynthesis and metabolism were concurrent with the anthocyanin content. The developmental stage appeared to affect the expression of genes related to flavonoid biosynthesis to a greater extent than the tissue or cultivar type. This study provides an abundant data resource that will further our understanding of the molecular mechanisms of anthocyanin biosynthesis in blueberries.

High-performance liquid chromatography for the analytical characterization of anthocyanins in Vaccinium myrtillus L. (bilberry) fruit and food products

Anthocyanins represent the most abundant class of bioactive compounds present in Vaccinium myrtillus L. (bilberry) fruit, conferring it several health-promoting properties. The content of anthocyanins in food products produced from bilberries can be affected by many parameters, making the study of their composition a critical issue. In this ambit, this work was aimed at a comprehensive profiling of anthocyanins in bilberry fruit and derivatives from the Italian Northern Apennines, including jam, juice, and liqueur ("Mirtillino"). Anthocyanins were extracted from the jams by means of a dynamic maceration with acidified methanol, while juice and liqueurs were directly analyzed. The analysis of anthocyanins in the extracts was carried out by means of HPLC-UV/DAD, HPLC-ESI-MS, and MS2, under gradient elution. As a comparison, authentic bilberry fruits were analyzed. The total anthocyanin content was in the range 582.4-795.2mg/100g (FW) for the fruit, 2.3-234.5mg/100g for the jams, 109.2-2252.2mg/L for the juice, and 27.9-759.3mg/L for the liqueurs. To deeper investigate the anthocyanin profile of the liqueurs that exhibited a remarkably different composition in comparison with the other products, an authentic bilberry liqueur was prepared in the lab, following a traditional recipe, and monitored weakly by HPLC. The percentage of degradation of 3-O-galactosides and 3-O-arabinosides of bilberry anthocyanidins was found to be higher than that of 3-O-glucosides. The results of this work demonstrated the importance of a suitable and reliable analysis of bilberry fruit and related food products to ensure their genuineness and quality. Graphical abstract Vaccinium myrtillus L. (bilberry) fruit and food products analyzed in this work.

Identification of basic/helix-loop-helix transcription factors reveals candidate genes involved in anthocyanin biosynthesis from the strawberry white-flesh mutant

As the second largest transcription factor family in plant, the basic helix-loop-helix (bHLH) transcription factor family, characterized by the conserved bHLH domain, plays a central regulatory role in many biological process. However, the bHLH transcription factor family of strawberry has not been systematically identified, especially for the anthocyanin biosynthesis. Here, we identified a total of 113 bHLH transcription factors and described their chromosomal distribution and bioinformatics for the diploid woodland strawberry Fragaria vesca. In addition, transcription profiles of 113 orthologous bHLH genes from various tissues were analyzed for the cultivar ‘Benihoppe’, its white-flesh mutant ‘Xiaobai’, and the ‘Snow Princess’ from their fruit development to the ripening, as well as those under either the ABA or Eth treatment. Both the RT-PCR and qRT-PCR results show that seven selected FabHLH genes (FabHLH17, FabHLH25, FabHLH27, FabHLH29, FabHLH40, FabHLH80, FabHLH98) are responsive to the fruit anthocyanin biosynthesis and hormone signaling according to transcript profiles where three color modes are observed for strawberry’s fruit skin and flesh. Further, prediction for the protein interaction network reveals that four bHLHs (FabHLH25, FabHLH29, FabHLH80, FabHLH98) are involved in the fruit anthocyanin biosynthesis and hormone signaling transduction. These bioinformatics and expression profiles provide a good basis for a further investigation of strawberry bHLH genes.

Anthocyanin Management in Fruits by Fertilization.

Anthocyanins are water-soluble vacuolar plant pigments that are mainly synthesized in epidermal layers and the flesh of fruits such as apples, cherries, grapes, and other berries. Because of their attractive red to purple coloration and their health-promoting potential, anthocyanins are significant determinants for the quality and market value of fruits and fruit-derived products. In crops, anthocyanin accumulation in leaves can be caused by nutrient deficiency which is usually ascribed to insufficient nitrogen or phosphorus fertilization. However, it is a little-known fact that the plant's nutrient status also impacts anthocyanin synthesis in fruits. Hence, strategic nutrient supply can be a powerful tool to modify the anthocyanin content and consequently the quality and market value of important agricultural commodities. Here we summarize the current knowledge of the influence of plant nutrients on anthocyanin synthesis in fruits of major global market value and discuss the underlying cellular processes that integrate nutrient signaling with fruit anthocyanin formation. It is highlighted that fertilization that is finely tuned in amount and timing has the potential to positively influence the fruit quality by regulating anthocyanin levels. We outline new approaches to enrich plant based foods with health-promoting anthocyanins.

FaTT12-1 , a multidrug and toxin extrusion (MATE) member involved in proanthocyanidin transport in strawberry fruits

Abstract Proanthocyanidin and anthocyanin comprise the most important phenolic components in strawberry fruits. Biosynthesis of these products has been extensively studied, but the transportation and trafficking processes are far from clear. Members of multidrug and toxin extrusion (MATE) transporters are imparted in the secondary metabolites transportation. In this study, we identified 53 MATE family members in the diploid Fragaria vesca genome. Based on homologous strategies, we obtained a potential AtTT12 orthologue in the octoploid strawberry, nominated as FaTT12-1. It could be detected in both vegetative and reproductive organs, but mainly in young tissues including young leaves and green fruits. Diverse cis-regulatory elements were recognized in the FaTT12-1 promoter. Blue light irradiation and abscisic acid treatment did not impact the expression of FaTT12-1. But red light irradiation can significantly elevate the transcription. Perturbation of FaTT12-1 via virus induced gene silencing (VIGS) in strawberry fruits did not give obvious visual changes. Fruit anthocyanin remained but proanthocyanidins were affected. In conclusion, we provide evidences that FaTT12-1 is a member of MATE family, specifically involved in proanthocyanidin accumulation in strawberry fruits.

A putative R3 MYB repressor is the candidate gene underlying atroviolacium, a locus for anthocyanin pigmentation in tomato fruit.

Anthocyanins are potential health-promoting compounds in the human diet. The atv (atroviolacium) locus, derived from the wild tomato species Solanum cheesmaniae, has been shown to enhance anthocyanin pigmentation in tomato fruit when it co-exists with either the Aft (Anthocyanin fruit) or the Abg (Aubergine) locus. In the present study, the atv locus was fine-mapped to an approximately 5.0-kb interval on chromosome 7. A putative R3 MYB repressor was identified in this interval and is hereby designated as SlMYBATV. The allele of SlMYBATV underlying the atv locus harbored a 4-bp insertion in its coding region, which is predicted to result in a frame-shift and premature protein truncation. The other candidate R3 MYB and R2R3 MYB repressors of anthocyanin biosynthesis were also identified in tomato via a genome-wide search. Transcriptional analysis showed that most of the structural genes and several regulatory genes of anthocyanin biosynthesis were up-regulated in the tomato SlMYBATV mutant lines. These findings may facilitate the elucidation of the molecular mechanisms underlying anthocyanin pigmentation in tomato fruit and help in the marker-assisted selection of anthocyanin-enriched tomato cultivars.

Abscisic acid stimulates anthocyanin accumulation in ‘Jersey’ highbush blueberry fruits during ripening

Non-climacteric blueberry (Vaccinium spp.) fruits accumulate high levels of anthocyanins during ripening, which are a good source of dietary antioxidants. This study examined the effects of exogenous abscisic acid (ABA) application on fruit characteristics and anthocyanin accumulation in a northern highbush blueberry (V. corymbosum ‘Jersey’) during development. Fruits on shrubs were treated with 1gL−1 ABA before the initiation of fruit colouration. Application of ABA temporarily increased the level of ABA in the fruits during development. Exogenous ABA had no obvious effect on fruit growth, but stimulated fruit colouration by accelerating the accumulation of individual anthocyanins, mainly malvidin, delphinidin and petunidin glycosides. This is the first report to show that ABA promotes the accumulation of anthocyanins in blueberry fruits. However, exogenous ABA also promoted fruit softening, which is undesirable during harvest and shelf life.

Protective effect of mulberry fruit anthocyanin on human hepatocyte cells (LO2) and Caenorhabditis elegans under hyperglycemic conditions

Type 2 diabetes is a chronic disease and hyperglycemia is important in the pathogenesis of diabetic complications. Exposure of LO2 cells to high glucose resulted in cellular glucose consumption and uptake decreases, reactive oxygen species (ROS) and superoxide anion (O2−) accumulation and mitochondrial dysfunction, which could be partly recovered by mulberry anthocyanin extract (MAE). And these protective effects were partly associated with regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream targets. As the insulin-signaling pathway is evolutionarily well conserved from Caenorhabditis elegans to mammals, C. elegans has been considered as a model system to study effects of glucose toxicity. Glucose shortened the lifespan of C. elegans, while MAE suppressed the damage, accompanied by malondialdehyde (MDA) and triglyceride accumulation reduction as well as total superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity recovery and PMK-1/p38 expression promotion. In contrast, MAE failed to recover shortened longevity, glucose and triglyceride accumulation in daf-2 (−) mutants fed a glucose-supplemented diet. Transcriptional profile revealed MAE intervention led to 92 genes alteration compared with the glucose-treatment. Interestingly, expressions of DAF-2/insulin receptor related genes were increased by glucose but impaired by MAE in nematodes. Our studies suggested that MAE might help to improve the antioxidant defense system, resulting in prevention of glucose-induced damage both in vitro and in vivo.

Season, location and cultivar influence on bioactive compounds of sour cherry fruits

The aim of this study was to determine how different locations, years and cultivars influenced polyphenol and anthocyanin content in fruits of different sour cherry cultivars (Prunus cerasus L.). Fruits of five sour cherry cultivars were harvested in two locations (Osijek and Zadar) over three consecutive years (2010, 2011 and 2012). Factorial ANOVA showed that year and location significantly influenced the accumulation of polyphenols and anthocyanins in sour cherry fruits. 2010 was the best year with 9.89 mg/g of polyphenols and 5.08 mg/g of anthocyanins on average. Although year and location revealed a strong influence, cultivar is the principal source of variation, as it is proved by the polyphenol content in the range from 5.89 to 10.78 mg/g and the anthocyanin content in the range from 3.15 to 4.76 mg/g. Cv. Maraska appears to be the most significant source of bioactive compounds, while cvs. Heimanns Konservenweichel and Rexelle gave very similar but significantly lower contents of polyphenols and anthocyanins than cv. Maraska. Cv. Oblačinska had significantly the lowest contents of investigated bioactive compounds.

Map-based cloning of the pear gene MYB114 identifies an interaction with other transcription factors to coordinately regulate fruit anthocyanin biosynthesis.

Red fruits are popular and widely accepted by consumers because of an enhanced appearance and enriched anthocyanins. The molecular mechanism of anthocyanin regulation in red-skinned pear (Pyrus) has been studied, and the genes encoding the biosynthetic steps and several transcription factors (TFs) have been characterized. In this study, a candidate R2R3 MYB TF, PyMYB114, was identified by linkage to the quantitative trait loci (QTL) for red skin color on linkage group 5 in a population of Chinese pear (Pyrus bretschneideri). The function of PyMYB114 was verified by transient transformation in tobacco (Nicotinana tabacum) leaves and strawberry (Fragaria) and pear fruits, resulting in the biosynthesis of anthocyanin. Suppression of PyMYB114 could inhibit anthocyanin biosynthesis in red-skinned pears. The ERF/AP2 TF PyERF3 was found to interact with PyMYB114 and its partner PybHLH3 to co-regulate anthocyanin biosynthesis, as shown by a dual luciferase reporter system and a yeast two-hybrid assay. In addition, the transcript abundance of PyMYB114 and PyMYB10 were correlated, and co-transformation of these two genes into tobacco and strawberry led to enhanced anthocyanin biosynthesis. This interaction network provides insight into the coloration of fruits and the interaction of different TFs to regulate anthocyanin biosynthesis.

Cloning and expression profiling of the PacSnRK2 and PacPP2C gene families during fruit development, ABA treatment, and dehydration stress in sweet cherry

Plant SNF1-related protein kinase 2 (SnRK2) and protein phosphatase 2C (PP2C) family members are core components of the ABA signal transduction pathway. SnRK2 and PP2C proteins have been suggested to play crucial roles in fruit ripening and improving plant tolerance to drought stress, but supporting genetic information has been lacking in sweet cherry (Prunus avium L.). Here, we cloned six full-length SnRK2 genes and three full-length PP2C genes from sweet cherry cv. Hong Deng. Quantitative PCR analysis revealed that PacSnRK2.2, PacSnRK2.3, PacSnRK2.6, and PacPP2C1–3 were negatively regulated in fruits in response to exogenous ABA treatment, PacSnRK2.4 and PacSnRK2.5 were upregulated, and PacSnRK2.1 expression was not affected. The ABA treatment also significantly promoted the accumulation of anthocyanins in sweet cherry fruit. The expression of all PacSnRK2 and PacPP2C genes was induced by dehydration stress, which also promoted the accumulation of drought stress signaling molecules in the sweet cherry fruits, including ABA, soluble sugars, and anthocyanin. Furthermore, a yeast two-hybrid analysis demonstrated that PacPP2C1 interacts with all six PacSnRK2s, while PacPP2C3 does not interact with PacSnRK2.5. PacPP2C2 does not interact with PacSnRK2.1 or PacSnRK2.4. These results indicate that PacSnRK2s and PacPP2Cs may play a variety of roles in the sweet cherry ABA signaling pathway and the fruit response to drought stress.