Physiological And Molecular Mechanisms Of Seedling Growth And Leaf Color Differences In Red-Leaved Poplar

Populus deltoides‘Quanhong’ and Populus deltoids ‘Xuanhong’ are bud sport speices of excellent variety Populus deltoides cv.‘Zhonglin2025’(Zhonglin 2025)with colored leaves.The leaves of Populus deltoides‘Quanhong’ are purple-red in spring,purple-green in summer and orange-yellow in autumn.The leaf color of Populus deltoides‘Xuanhong’ is more bright and persistent than that of Populus deltoides‘Quanhong’,and the leaf color gradually changes from bright red at budding stage to orange red or orange yellow at deciduous stage.They all have the excellent characteristics of beautiful color and beautiful tree shape,and have broad application prospects in landscaping.However,in the production and application of seedlings it was found that the growth potential of Populus deltoides‘Quanhong’ and Populus deltoides‘Xuanhong’ was significantly lower than that of Zhonglin 2025.In order to reveal the physiological and molecular mechanism of differences in leaf color and growth between red leaf poplar and Zhonglin 2025,the 1-year-old seedlings of Populus deltoides‘Quanhong’ and Populus deltoides ‘Xuanhong’ were used as test materials,and the 1-year-old seedlings of Zhonglin 2025 were used as control.The physiological indexes such as phenotypic characteristics,photosynthesis parameters and chlorophyll fluorescence parameters were measured to clarify the physiological reasons for the inhibition of leaf color formation and growth of red leaf poplar;through transcriptome sequencing and protein sequencing,key genes and proteins controlling leaf color formation and photosynthetic capacity decline were screened out.The main results are as follows:(1)The growth amount of Populus deltoides ‘xuanhong’ and Populus deltoides‘Quanhong’ was significantly lower than that of Zhonglin 2025(p<0.01).The height growth of 1-year-old seedlings is 17.95% and 32.79% of that of Zhonglin 2025,respectively.The ground diameter growth is 32.49% and 57.63% of that of Zhonglin 2025,respectively,The number of leaves was 28.81% and 29.22% of that of Zhonglin 2025,and the leaf area was39.68% and 48.78% of Zhonglin 2025,respectively.(2)The anthocyanin content of Populus deltoides ‘xuanhong’ and Populus deltoides‘Quanhong’ was much higher than that of Zhonglin 2025;the chlorophyll a,chlorophyll b and total chlorophyll contents of the three poplar species were Populus deltoides ‘Quanhong’>Zhonglin 2025 > Populus deltoides ‘Xuanhong’,Populus deltoides ‘Xuanhong’was significantly lower than Populus deltoides ‘Quanhong’ and Zhonglin 2025.The leaves of Populus deltoides ‘Quanhong’ were red,and the ratio of anthocyanin to chlorophyll(Ant/Chl)was 12.9;the leaves of Populus deltoides ‘Quanhong’ were purple-red with a ratio of 2.7;Zhonglin 2025 was green with a ratio of 0.4.The higher anthocyanin content is one of the main reasons why Populus deltoides ‘Xuanhong’ and Populus deltoides ‘Quanhong’ show red color.The purple color of Populus deltoides ‘Quanhong’ leaves is due to the higher chlorophyll content and anthocyanin content in its leaves.(3)The net photosynthetic rate of the three poplar species in May,July and September showed that: Populus deltoides ‘Quanhong’ and Populus deltoides ‘Xuanhong’ were significantly lower than Zhonglin 2025;the Populus deltoides ‘Xuanhong’ was lower than the Populus deltoides ‘Quanhong’,but the difference between them was not significant.The diurnal variation of net photosynthetic rate of Zhonglin 2025 and Populus deltoides‘Quanhong’ in May and July was a bimodal curve,and there was a phenomenon of "midday rest" in photosynthesis.The diurnal variation of net photosynthetic rate of Populus deltoides‘Xuanhong’ was a unimodal curve.In September,the diurnal variation of net photosynthetic rate of the three poplar species was a unimodal curve.When "photosynthetic midday rest" appeared in Zhonglin 2025 and Populus deltoides ‘Quanhong’,Gs was in a declining state,while Ci showed an upward trend,suggesting that "photosynthetic midday rest" was mainly affected by non-stomatal constraints.(4)The Populus deltoides ‘Xuanhong’ has obvious reflection peaks in the red light region,and the absorption rate of red light is low,which affects its photosynthetic rate.There is no obvious spectral reflection peak in the visible range of Populus deltoides ‘Quanhong’,which has a strong absorption ability to visible light.Compared with Zhonglin 2025,Quantum yield Y(NPQ)of PSII regulatory energy dissipation of Populus deltoides‘Quanhong’ is larger,and actual photosynthetic efficiency(YII)of PSII of Populus deltoides‘Quanhong’ is lower,resulting in more heat dissipation energy of PSII of photosynthetic system,low actual light energy conversion efficiency and low photosynthetic capacity of Populus deltoides ‘Quanhong’.(5)Transcriptome sequencing of the leaves of two red-leaf poplars and Zhonglin 2025 showed that a total of about 58.6 G Clean bases were obtained,with an average amount of about 6.5 G per sample.The sequencing error rate,Q20 value,Q30 value and GC content of all samples were within reasonable range,and the proportion of successful sequencing reads of each sample compared to the reference genome of Populus tomentosa was higher than80%.Through differential screening,6792 and 4398 differentially expressed genes were obtained from Populus deltoides ‘Quanhong’ and Populus deltoides ‘Xuanhong’,respectively,and 2779 differentially expressed genes were shared by the two species.GO annotation and KEGG enrichment analysis results showed that the differentially expressed genes in Populus deltoides ‘Quanhong’ and Populus deltoides ‘Xuanhong’ were significantly enriched in 21 and 18 metabolic pathways,including carbon fixation,amino acid biosynthesis,secondary metabolite biosynthesis,metabolic pathway,carbon metabolism,glycolysis pathway,tyrosine metabolism,cysteine and methionine metabolism,isoquinoline alkaloids biosynthesis,flavonoid biosynthesis,and alpha-linolenic acid metabolic pathways.Compared with the two red-leaf poplar species,the differentially expressed genes in Populus deltoides ‘Quanhong’ species were significantly enriched in more Go terms and metabolic pathways.(6)2786 and 2333 differentially expressed proteins were screened from Populus deltoides‘Quanhong’and Populus deltoides‘Xuanhong’by TMT markers,respectively.GO and KEGG enrichment analysis showed that the differentially expressed proteins in Populus deltoides‘Quanhong’and Populus deltoides ‘Xuanhong’were significantly enriched in 48 and 26 metabolic pathways,respectively,and they shared 25 metabolic pathways including carbon metabolism,metabolic pathway and biosynthesis of secondary metabolites.(7)Joint analysis of transcriptome and proteome showed that 98.70% and 98.75%proteins were contained in the genes of Populus deltoides ‘Quanhong’ and Populus deltoides‘Xuanhong’ compared with Zhonglin 2025,respectively.Genes and proteins in Populus deltoides ‘Quanhong’ and Populus deltoides ‘Xuanhong’ were significantly enriched in important metabolic pathways such as photosynthesis,phenylpropane metabolism and flavonoid synthesis.In addition to the significant upregulation of Psb R gene,several chloroplast protein-coding genes were significantly downregulated in the photosynthesis pathway of Populus deltoides ‘Quanhong’,and the expression of more subunit proteins was significantly decreased in Populus deltoides ‘Xuanhong’,indicating that the electron transport and photophosphorylation processes of two species of red-leaf poplar were greatly affected,thereby reducing the photosynthetic capacity.(8)15 and 11 genes and proteins,including PAL,CHS,CHI,F3 H and F3’H,were identified from Populus deltoides ‘Quanhong’ and Populus deltoides ‘Xuanhong’ which participated in controlling the synthesis of anthocyanins.In the pathway of pigment metabolism,the expression of PAL,CHS,CH4,CHI,F3 H,F3’H and DFR genes and proteins controlling anthocyanin synthesis were significantly up-regulated in Populus deltoides‘Quanhong’;in Populus deltoides ‘Xuanhong’,the expression of PAL,CHS,CHI,F3 H,F3’H,DFR and ANS genes and proteins was significantly up-regulated.This indicates that the leaf color of red-leaf poplar is closely related to the up-regulated expression of several genes controlling anthocyanin structure.Based on the results of joint analysis,three transcription factors,TTG2,HYH and HY5,may play important regulatory roles in controlling anthocyanin synthesis.