To assess the interrelation between the change of metabolites and the change of fruit color, we performed a combined metabolome and transcriptome analysis of the flesh in two different Actinidia arguta cultivars: “HB” (“Hongbaoshixing”) and “YF” (“Yongfengyihao”) at two different fruit developmental stages: 70d (days after full bloom) and 100d (days after full bloom). Metabolite and transcript profiling was obtained by ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometer and high-throughput RNA sequencing, respectively. The identification and quantification results of metabolites showed that a total of 28,837 metabolites had been obtained, of which 13,715 were annotated. In comparison of HB100 vs. HB70, 41 metabolites were identified as being flavonoids, 7 of which, with significant difference, were identified as bracteatin, luteolin, dihydromyricetin, cyanidin, pelargonidin, delphinidin and (−)-epigallocatechin. Association analysis between metabolome and transcriptome revealed that there were two metabolic pathways presenting significant differences during fruit development, one of which was flavonoid biosynthesis, in which 14 structural genes were selected to conduct expression analysis, as well as 5 transcription factor genes obtained by transcriptome analysis. RT-qPCR results and cluster analysis revealed that AaF3H, AaLDOX, AaUFGT, AaMYB, AabHLH, and AaHB2 showed the best possibility of being candidate genes. A regulatory network of flavonoid biosynthesis was established to illustrate differentially expressed candidate genes involved in accumulation of metabolites with significant differences, inducing red coloring during fruit development. Such a regulatory network linking genes and flavonoids revealed a system involved in the pigmentation of all-red-fleshed and all-green-fleshed A. arguta, suggesting this conjunct analysis approach is not only useful in understanding the relationship between genotype and phenotype, but is also a powerful tool for providing more valuable information for breeding.

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Cold stress poses a serious treat to cultivated kiwifruit since this plant generally has a weak ability to tolerate freezing tolerance temperatures. Surprisingly, however, the underlying mechanism of kiwifruit’s freezing tolerance remains largely unexplored and unknown, especially regarding the key pathways involved in conferring this key tolerance trait. Here, we studied the metabolome and transcriptome profiles of the freezing-tolerant genotype KL (Actinidia arguta) and freezing-sensitive genotype RB (A. arguta), to identify the main pathways and important metabolites related to their freezing tolerance. A total of 565 metabolites were detected by a wide-targeting metabolomics method. Under (−25°C) cold stress, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway annotations showed that the flavonoid metabolic pathways were specifically upregulated in KL, which increased its ability to scavenge for reactive oxygen species (ROS). The transcriptome changes identified in KL were accompanied by the specific upregulation of a codeinone reductase gene, a chalcone isomerase gene, and an anthocyanin 5-aromatic acyltransferase gene. Nucleotides metabolism and phenolic acids metabolism pathways were specifically upregulated in RB, which indicated that RB had a higher energy metabolism and weaker dormancy ability. Since the LPCs (LysoPC), LPEs (LysoPE) and free fatty acids were accumulated simultaneously in both genotypes, these could serve as biomarkers of cold-induced frost damages. These key metabolism components evidently participated in the regulation of freezing tolerance of both kiwifruit genotypes. In conclusion, the results of this study demonstrated the inherent differences in the composition and activity of metabolites between KL and RB under cold stress conditions.

As important regulators, miRNAs could play pivotal roles in regulation of fruit coloring. Actinidia arguta is a newly emerged fruit tree with extensively application prospects. However, miRNAs involved in A . arguta fruit coloring are unknown. In this study, A . arguta fruit were investigated at three developmental stages by small RNAs high-throughput sequencing. A total of 482 conserved miRNAs corresponding to 526 pre-miRNAs and 581 novel miRNAs corresponding to 619 pre-miRNAs were grouped into 46 miRNA families. Target gene prediction and analysis revealed that miR858, a strongly candidate miRNA, was involved in anthocyanin biosynthesis in which contributes to fruit coloring. The anthocyanin level was determined in three A . arguta cultivars by UPLC-MS/MS (ultra-performance liquid chromatography coupled with tandem mass spectrometry). In addition, qPCR (quantitative real-time PCR), cluster analysis were conducted as well as correlation analysis. All results were combined to propose a model in which describes an association of miRNA and anthocyanin biosynthesis in A . arguta . The data presented herein is the first report on miRNA profile analysis in A . arguta , which can provide valuable information for further research into the regulation of the miRNAs in anthocyanin biosynthesis and fruit coloring.

Actinidia arguta ‘Tianyuanhong’ is a new kiwifruit variety with an all-red pericarp and pulp, in contrast to the all-green pulp of A. arguta ‘Yongfengyihao’. Transcriptome profile analysis of fruit color has been reported, however, the metabolic mechanisms producing red flesh remain unknown, and it is unclear why the pulp of ‘Tianyuanhong’ is red rather than green. Herein, we identified differences between the proteomes of two A. arguta cultivars with different fruit color by using iTRAQ-based quantitative proteomic methods during the stage of color change. In total, 2310 differentially abundant proteins were detected between the two cultivars at 70 and 100 days after flowering, and the protein functions were analyzed based on KEGG and GO. The largest group of differentially expressed proteins were related to photosynthesis, glyoxylate metabolism, N metabolism, and anthocyanin biosynthesis. Finally, to verify the iTRAQ data, 12 representative genes encoding differentially expressed proteins were analyzed via quantitative real-time PCR, and these genes differed in transcriptional and translational expression levels. Our proteomic study contributes to understanding the metabolic pathways and biological processes involved in fruit color changes in different cultivars of A. arguta. These data and analyses will provide new insight into the development of kiwifruit flesh color.

Low temperature causes injuries to plants during winter, thereby it affects kiwi fruit quality and yield. However, the changes in metabolites and gene expression during cold acclimation (CA) and deacclimation (DA) in kiwi fruit remain largely unknown. In this study, freezing tolerance, carbohydrate metabolism, and β-amylase gene expression in two Actinidia arguta cv. “CJ-1” and “RB-3” were detected from CA to DA stages. In all acclimation stages, the “CJ-1” was hardier than “RB-3” and possessed lower semi-lethal temperature (LT50). Furthermore, “CJ-1” had a more rapid acclimation speed than “RB-3”. Changes of starch, β-amylase, and soluble sugars were associated with freezing tolerance in both cultivars. Starch contents continued to follow a declining trend, while soluble sugars contents continuously accumulated in both cultivars during CA stages (from October to January). To investigate the possible molecular mechanism underlying cold response in A. arguta, in total, 16 AcBAMs genes for β-amylase were identified in the kiwi fruit genome. We carried out localization of chromosome, gene structure, the conserved motif, and the analysis of events in the duplication of genes from AcBAMs. Finally, a strong candidate gene named AaBAM3 from AcBAMs was cloned in Actinidia arguta (A. arguta), The real-time qPCR showed that AaBAM3 gene expression in seasonal changes was consistent with changes of soluble sugars. These results reveal that AaBAM3 may enhance the freezing tolerance of A. arguta through increasing soluble sugar content.

Background Freezing injury, which is an important abiotic stress in horticultural crops, influences the growth and development and the production area of kiwifruit (Actinidia Lind1). Among Actinidia species, Actinidia arguta has excellent cold resistance, but knowledge relevant to molecular mechanisms is still limited. Understanding the mechanism underlying cold resistance in kiwifruit is important for breeding cold resistance. Results In our study, a population resulting from the cross of A. arguta ‘Ruby-3’ × ‘Kuilv’ male was generated for kiwifruit hardiness study, and 20 cold-tolerant and 20 cold-sensitive populations were selected from 492 populations according to their LT50. Then, we performed bulked segregant RNA-seq combined with single-molecule real-time sequencing to identify differentially expressed genes that provide cold hardiness. We found that the content of soluble sucrose and the activity of β-amylase were higher in the cold-tolerant population than in the cold-sensitive population. Upon − 30 °C low-temperature treatment, 126 differentially expressed genes were identify; the expression of 59 genes was up-regulated and that of 67 genes was down-regulated between the tolerant and sensitive pools, respectively. KEGG pathway analysis showed that the DEGs were primarily related to starch and sucrose metabolism, amino sugar and nucleotide sugar metabolism. Ten major key enzyme-encoding genes and two regulatory genes were up-regulated in the tolerant pool, and regulatory genes of the CBF pathway were found to be differentially expressed. In particular, a 14–3-3 gene was down-regulated and an EBF gene was up-regulated. To validate the BSR-Seq results, 24 DEGs were assessed via qRT-PCR, and the results were consistent with those obtained by BSR-Seq. Conclusion Our research provides valuable insights into the mechanism related to cold resistance in Actinidia and identified potential genes that are important for cold resistance in kiwifruit.