Transcriptome and metabolome combined analysis of anthocyanin metabolism in fruit peel of Ziziphus jujuba Mill.‘Lingwuchangzao’under the interaction of elevated temperature and drought

Abstract: 【Objective】Ziziphus jujuba Mill.‘Lingwuchangzao'is an excellent fresh jujube variety in Ningxia. Global warming has become the main feature of global climate change in the recent 100 years,while climate warming leads to changes in precipitation. Elevated air temperature and drought are two important environmental factors influencing fruit coloring and quality. Therefore, key genes regulating fruit color under the synergistic treatment of elevated air temperature and drought were screened, and understanding related metabolism provide important reference for the response of fruit quality formation process of economic forest to climate change.【Methods】In this experiment, jujube cultivar Lingwuchangzao was used as the experimental object, and the combined treatment of air temperature and drought was applied. The open-top chamber control system was used to simulate the elevated temperature, automatic water-saving irrigation system to control soil moisture levels. The air temperature was set at two levels [natural environmental temperature T1, elevated air temperature T2=T1+(2.0±0.5 ℃)],and soil moisture was set to three levels(normal soil moisture D1 with a relative soil moisture content in the range of 70%-75% of field capacity; moderate drought D2 with a relative soil moisture content in the range of 50%-55% of field capacity; and severe drought D3 with a relative soil moisture content of30%-35% of field capacity). Totally there were 6 treatments, with natural air temperature and normal soil moisture serving as the control. Anthocyanin content in the peel at different developmental stages was determined, and the coloring differences between the treatments were compared. Peel samples were taken for transcription sequencing and metabolite determination. Key regulatory genes were selected and analyzed with qRT-PCR.【Results】With the gradual maturation of fruit, the anthocyanin content showed a gradual increasing trend under the synergistic treatment of elevated temperature and drought.When the air temperature rose by 2 ℃, the anthocyanin content increased by 6.59%-47.64%. Drought did not favor the accumulation of anthocyanin in the peel, and the anthocyanin content decreased by4.28%-12.33% under the combined treatment of elevated temperature and drought. A total of 129.66 Gb clean data were obtained by transcription sequencing and a total of 29 610 genes were detected with high FPKM values. The maximum number of differentially expressed genes was 6599 under the combined treatment of elevated temperature and severe drought at fully ripe stage. Based on MAPMAN annotation analysis, 6278 differentially expressed genes were annotated to the total metabolic pathway, and 8 genes(3 up-regulated and 5 down-regulated) were annotated in the flavonoid metabolism of the secondary metabolic pathway. KEGG enrichment analysis showed that there were 13 up-regulated differential genes in the comparison between severe drought and control, 3 down-regulated and 10 up-regulated in response to elevated temperature combined with severe drought treatment. It was noted that 8 key genes and seven differential metabolites were identified in the flavonoid metabolism. Pelargonidin, epicatechin and delphindin, the final products generated from the key metabolite cinnamoyl-CoA under the regulation of related genes and enzymes, were associated with the synthesis and transformation of anthocyanins. In this process, the eight key differential genes were significantly up-regulated, and the contents of the seven metabolites in different metabolic groups were different. Construction of the regulatory network between different expressed genes and metabolites showed that chalcone synthase(CHS), bifunctional dihydroflavonol 4-reductase(DFR), anthocyanidin synthase(ANS) and chalcone--flavonone isomerase(CFI) genes were significant up regulated under elevated temperatures and drought treatment, which promoted the accumulation of kaempferol, quercetin and pigment. The accumulation of the seven key metabolites was promoted under the treatment with elevated temperature and normal soil moisture, but decreased under elevated temperature combined with severe drought. q PCR analysis showed that the key regulatory genes were significantly up regulated in different treatment materials, and the expression trend was consistent with the transcription.【Conclusion】Elevated air temperature and drought had significant effects on fruit coloring of Lingwuchangzao, which could be well explained at both physiological and genetic levels. There was significant difference in anthocyanin content between treatments. The anthocyanin content was increased byevaluated temperature, and decreased by drought. Transcription data showed that 4 genes were identified from the Lingwuchangzao, namely CHS, DFR, ANS and CFI genes, which were expressed in fruit peel and the expression levels were different in each treatment.ZjCHS, ZjDFR, ZjANS and ZjCFI played an essential regulatory role in anthocyanin synthesis under the synergistic treatment of evaluated air temperature and drought in Lingwuchangzao. The results of this study will provide experimental support for subsequent functional studies.