To explore the mechanism underlying the leaf color variation of Populus × euramericana ‘Zhonghuahongye’ (‘Zhonghong’ poplar) leaves at different maturation stages, we used ‘Zhonghong’ poplar leaves and analyzed the L* (lightness), a* (redness), and b* (yellowness) color difference values and the pigment contents in the three maturation stages. The Illumina HiSeqTM 2000 high−throughput sequencing platform was used for transcriptome sequencing analysis, and leaf color changes during stage transitions were preliminarily explored. Overall, with the increase in L* and b* during leaf development, the a*, C* (colour saturation), and chromatic values decreased, the total anthocyanin content decreased, and the chlorophyll and carotenoid content increased. A total of 11,868 differentially expressed genes were identified by transcriptome sequencing. Comparing the expression differences of structural genes involved in anthocyanin synthesis in the leaves at different stages, we identified 5612 genes in the R1 vs. R2 comparison, 10,083 in the R1 vs. R3 comparison, and 6068 in the R2 vs. R3 comparison (R1, R2, R3 refer to samples obtained on 1 April, 6 April, and 11 April, respectively). Key genes such as DFR (Dihydroflavanol 4−reductase), ANS (anthocyanidin synthase), FLS(flavonol synthase) , CHS(chalcone synthase), BZ1(Bronze 1), bHLH35, and bHLH63 were identified. These structural genes and those that encode transcription factors may be related to the regulation of anthocyanin synthesis. Here, the key genes related to leaf color change in ‘Zhonghong’ poplar were discovered, providing an important genetic basis for the subsequent genetic improvement of ‘Zhonghong’ poplar.

Basic helix–loop–helix (bHLH) proteins belong to one of the largest families involved in plant growth, development, signal transduction, and secondary metabolism. Although bHLH genes have been previously identified in persimmon (Diospyros kaki), systematic studies have not been reported. A total of 59 bHLH family members have been identified from the “Xiaoguotianshi” persimmon transcriptome. These proteins were clustered into 12 groups from I to XII based on their phylogenetic relationships with Arabidopsis thaliana. Combined with the phylogenetic analysis, in silico expression patterns of five developmental stages, the protein–protein interaction analysis between DkbHLH and DkMYB proteins showed that the bHLH_Cluster-15548.1 protein sequence was identified to be highly similar to the AtGL3 (AT5G41315.1) protein, which is associated with flavonoid and proanthocyanidin (PA) biosynthesis. This study presents the systematic analysis of bHLH genes from D. kaki and provides valuable information for further research on the involvement of bHLH protein in anthocyanin biosynthesis.