Comparative Transcriptome and Metabolome Profiling Reveal Mechanisms of Red Leaf Color Fading in Populus × euramericana cv. 'Zhonghuahongye'

Abstract

Anthocyanins are among the flavonoids that serve as the principal pigments affecting the color of plants. During leaf growth, the leaf color of 'Zhonghuahongye' gradually changes from copper-brown to yellow-green. At present, the mechanism of color change at different stages has not yet been discovered. To find this, we compared the color phenotype, metabolome, and transcriptome of the three leaf stages. The results showed that the anthocyanin content of leaves decreased by 62.5% and the chlorophyll content increased by 204.35%, 69.23%, 155.56% and 60%, respectively. Differential metabolites and genes were enriched in the pathway related to the synthesis of 'Zhonghuahongye' flavonoids and anthocyanins and to the biosynthesis of secondary metabolites. Furthermore, 273 flavonoid metabolites were detected, with a total of eight classes. DFR, FLS and ANS downstream of anthocyanin synthesis may be the key structural genes in reducing anthocyanin synthesis and accumulation in the green leaf of 'Zhonghuahongye'. The results of multi-omics analysis showed that the formation of color was primarily affected by anthocyanin regulation and its related synthesis-affected genes. This study preliminarily analyzed the green regression gene and metabolic changes in 'Zhonghuahongye' red leaves and constitutes a reference for the molecular breeding of 'Zhonghuahongye' red leaves.

Keywords: anthocyanin; metabolome; molecular breeding; the color fading of red leaves; transcriptome.

Figure 1

Comparison of leaf color and phenotype changes: (a) leaf phenotypic color determination at different periods (scale bar: 1cm); (b) microstructure of leaves at different periods(scale bar: Petiole: 200 μm; Vein: 200 μm; Mescophyll: 100 μm). Note: G1: 1 April; G2: 6 April; G3: 11 April.

Figure 2

Qualitative and quantitative analysis of leaf metabolites in different periods: (a) sample aggregate metabolite cluster diagram; (b) PCA score chart of quality spectrum data of samples and quality control samples for each group; (c) sample aggregate metabolite clustering diagram.

Figure 3

Analysis of leaf metabolites difference in different periods: (a) the amount of leaf metabolites in different periods; (b) differential metabolite volcano map; (c) Venn diagram; (d) histogram of metabolite difference multiples.

Figure 4

Leaf transcriptome analysis of ‘Zhonghuahongye’ at different periods: (a) differential gene statistical map; (b) differential gene volcano map; (c) k-means cluster graph; (d) correlation heat map; (e) Venn diagram; (f) differential gene KEGG enrichment map.

Figure 5

Nine-quadrant diagram of correlation analysis. Note: The horizontal coordinate indicates the log2FC of the gene and the vertical coordinate indicates the log2FC of the metabolite.

Figure 6

Correlation clustering heatmap. Note: In the figure, each row has a gene, each column is a metabolite, red represents a positive correlation between the gene and metabolite, and green represents a negative correlation between the gene and the metabolite.

Figure 7

CCA diagram. Note: In the figure, four regions are distinguished by crosses. In the same region, the farther away from the origin a point was, the closer the distance between each other and the higher the correlation. Metabolites are labeled purple and genes are labeled red. If there is too much of a certain type of substance, in order to avoid overlapping text, it will be displayed with dots instead.

Figure 8

Bioanabolism of anthocyanins in the leaves of ‘Zhonghuahongye’. Note: Each color circle represents the level of gene expression according to the color scale. The enzymes bioanabolically associated with anthocyanins of ‘Zhonghuahongye’ are shown in Table S3. Detailed information for 83 differentially expressed genes is shown in Table S4.