Transcriptome-wide m6A methylation in natural yellow leaf of Catalpa fargesii.

N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNA, and involved in various biological processes in plants. However, the distribution features and functions of mRNA m6A methylation have been poorly explored in woody perennial plants. In this study, a new natural variety with yellow-green leaves, named Maiyuanjinqiu, was screened from the seedlings of Catalpa fargesii. Based on the preliminary experiment, the m6A methylation levels in the leaves of Maiyuanjinqiu were significantly higher than those in C. fargesii. Furthermore, a parallel analysis of m6A-seq and RNA-seq was carried out in different leaf color sectors. The result showed that m6A modification were mostly identified around the 3'-untranslated regions (3'-UTR), which was slightly negatively correlated with the mRNA abundance. KEGG and GO analyses showed that m6A methylation genes were associated with photosynthesis, pigments biosynthesis and metabolism, oxidation-reduction and response to stress, etc. The overall increase of m6A methylation levels in yellow-green leaves might be associated with the decreased the expression of RNA demethylase gene CfALKBH5. The silencing of CfALKBH5 caused a chlorotic phenotype and increased m6A methylation level, which further confirmed our hypothesis. Our results suggested that mRNA m6A methylation could be considered as a vital epigenomic mark and contribute to the natural variations in plants.

Genetic architecture of leaf morphology revealed by integrated trait module in Catalpa bungei.

Leaves are crucial for maintaining plant growth and development via photosynthesis, and their function is simultaneously regulated by a suite of phenotypic traits. Although much is known about the genetic architecture of individual leaf traits, unraveling the genetic basis of complex leaf morphology remains a challenge. Based on the functional correlation and coordination of multi-traits, we divided 15 leaf morphological traits into three modules, comprising size (area, length, width, and perimeter), shape (leaf lobes, aspect ratio, circularity, rectangularity, and the relevant ratios), and color (red, green, and blue) for an ornamental tree species, Catalpa bungei. A total of 189 significant single-nucleotide polymorphisms were identified in the leaves of C. bungei: 35, 82, and 76 in the size, shape, and color modules, respectively. Four quantitative trait loci were common between the size and shape modules, which were closely related according to phenotype correlation, genetic mapping, and mRNA analysis. The color module was independent of them. Synergistic changes in the aspect ratio, leaf lobe, and circularity suggest that these traits could be the core indicators of the leaf shape module. The LAS and SRK genes, associated with leaf lobe and circularity, were found to function in plant defense mechanisms and the growth of leaves. The associations between the SRK and CRK2 genes and the leaf lobe and circularity traits were further verified by RT-qPCR. Our findings demonstrate the importance of integrating multi-trait modules to characterize leaf morphology and facilitate a holistic understanding of the genetic architecture of intraspecific leaf morphology diversity.

Genome-wide identification of DnaJ gene family in Catalpa bungei and functional analysis of CbuDnaJ49 in leaf color formation.

DnaJs are the common molecular chaperone proteins with strong structural and functional diversity. In recent years, only several DnaJ family members have been found to be able to regulate leaf color, and it remains to be explored whether there are other potential members that also regulate this character. Here, we identified 88 putative DnaJ proteins from Catalpa bungei, and classified them into four types according to their domain. Gene-structure analysis revealed that each member of CbuDnaJ family had same or similar exon-intron structure. Chromosome mapping and collinearity analysis showed that tandem and fragment duplication occurred in the process of evolution. Promoter analyses suggested that CbuDnaJs might be involved in a variety of biological processes. The expression levels of DnaJ family members in different color leaves of Maiyuanjinqiu were respectively extracted from the differential transcriptome. Among these, CbuDnaJ49 was the largest differentially expressed gene between the green and yellow sectors. Ectopic overexpression of CbuDnaJ49 in tobacco showed that the positive transgenic seedlings exhibited albino leaves, and the contents of chlorophyll and carotenoid were significantly reduced compared with those of wild type. The results suggested that CbuDnaJ49 played an important role in regulating leaf color. This study not only identified a novel gene of DnaJ family members regulating leaf color, but also provided new germplasm for landscaping.