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Background Liriodendron chinense is a distinctive ornamental tree species due to its unique leaves and tulip-like flowers. The discovery of genes involved in leaf development and morphogenesis is critical for uncovering the underlying genetic basis of these traits. Genes in the AP2/ERF family are recognized as plant-specific transcription factors that contribute to plant growth, hormone-induced development, ethylene response factors, and stress responses. Results In this study, we identified 104 putative AP2/ERF genes in the recently released L. chinense genome and transcriptome database. In addition, all 104 genes were grouped into four subfamilies, the AP2, ERF, RAV, and Soloist subfamilies. This classification was further supported by the results of gene structure and conserved motif analyses. Intriguingly, after application of a series test of cluster analysis, three AP2 genes, LcERF 94, LcERF 96, and LcERF 98, were identified as tissue-specific in buds based on the expression profiles of various tissues. These results were further validated via RT-qPCR assays and were highly consistent with the STC analysis. We further investigated the dynamic changes of immature leaves by dissecting fresh shoots into seven discontinuous periods, which were empirically identified as shoot apical meristem (SAM), leaf primordia and tender leaf developmental stages according to the anatomic structure. Subsequently, these three candidates were highly expressed in SAM and leaf primordia but rarely in tender leaves, indicating that they were mainly involved in early leaf development and morphogenesis. Moreover, these three genes displayed nuclear subcellular localizations through the transient transformation of tobacco epidermal cells. Conclusions Overall, we identified 104 AP2/ERF family members at the genome-wide level and discerned three candidate genes that might participate in the development and morphogenesis of the leaf primordium in L. chinense.

The MYB transcription factor family is one of the largest families in plants, and its members have various biological functions. R2R3-MYB genes are involved in the synthesis of pigments that yield petal colors. Liriodendron plants are widely cultivated as ornamental trees owing to their peculiar leaves, tulip-like flowers, and colorful petals. However, the mechanism underlying petal coloring in this species is unknown, and minimal information about MYB genes in Liriodendron is available. Herein, this study aimed to discern gene(s) involved in petal coloration in Liriodendron via genome-wide identification, HPLC, and RT-qPCR assays. In total, 204 LcMYB superfamily genes were identified in the Liriodendron chinense genome, and 85 R2R3-MYB genes were mapped onto 19 chromosomes. Chromosome 4 contained the most (10) R2R3-MYB genes, and chromosomes 14 and 16 contained the fewest (only one). MEME analysis showed that R2R3-MYB proteins in L. chinense were highly conserved and that their exon-intron structures varied. The HPLC results showed that three major carotenoids were uniformly distributed in the petals of L. chinense, while lycopene and β-carotene were concentrated in the orange band region in the petals of Liriodendron tulipifera. Furthermore, the expression profiles via RT-qPCR assays revealed that four R2R3-MYB genes were expressed at the highest levels at the S3P/S4P stage in L. tulipifera. This result combined with the HPLC results showed that these four R2R3-MYB genes might participate in carotenoid synthesis in the petals of L. tulipifera. This work laid a cornerstone for further functional characterization of R2R3-MYB genes in Liriodendron plants.

The MYB transcription factor family is one of the largest families in plants, and its members have various biological functions. R2R3-MYB genes are involved in the synthesis of pigments that yield petal colors. Liriodendron plants are widely cultivated as ornamental trees owing to their peculiar leaves, tulip-like flowers, and colorful petals. However, the mechanism underlying petal coloring in this species is unknown, and minimal information about MYB genes in Liriodendron is available. Herein, this study aimed to discern gene(s) involved in petal coloration in Liriodendron via genome-wide identification, HPLC, and RT-qPCR assays. In total, 204 LcMYB superfamily genes were identified in the Liriodendron chinense genome, and 85 R2R3-MYB genes were mapped onto 19 chromosomes. Chromosome 4 contained the most (10) R2R3-MYB genes, and chromosomes 14 and 16 contained the fewest (only one). MEME analysis showed that R2R3-MYB proteins in L. chinense were highly conserved and that their exon-intron structures varied. The HPLC results showed that three major carotenoids were uniformly distributed in the petals of L. chinense, while lycopene and β-carotene were concentrated in the orange band region in the petals of Liriodendron tulipifera. Furthermore, the expression profiles via RT-qPCR assays revealed that four R2R3-MYB genes were expressed at the highest levels at the S3P/S4P stage in L. tulipifera. This result combined with the HPLC results showed that these four R2R3-MYB genes might participate in carotenoid synthesis in the petals of L. tulipifera. This work laid a cornerstone for further functional characterization of R2R3-MYB genes in Liriodendron plants.

Floral fragrance, which has the function of attracting pollinators, is a class of volatile secondary metabolites mainly released by the secretory tissue of petals. Terpenoids are key components of floral volatile substances. Previous studies have shown that there are significant differences in the concentration and composition of volatile floral fragrances, especially terpenoids, between Liriodendron chinense and L. tulipifera. At present, the mechanism by which the synthesis of floral fragrance is regulated in Liriodendron remains unexplored. In this study, we analyzed the differentially expressed genes (DEGs) of L. chinense and L. tulipifera, and identified 130 DEGs related to terpenoid synthesis. A KEGG enrichment analysis of DEGs related to terpenoid biosynthesis revealed that the monoterpenoid biosynthesis pathway was the most significant. We cloned the LtuDXR gene from L. tulipifera using RACE technology. RT-qPCR results showed that the expression of the LtuDXR gene was the highest in the early florescence petals, indicating that the LtuDXR gene may play a role in the synthesis of volatile terpenoids. Subcellular localization showed that the LtuDXR protein is mainly localized in the chloroplast. Overexpression of LtuDXR in Arabidopsis thaliana significantly increased the plant height, DXR enzyme activity, and carotenoid content. In this study, we identified and functionally characterized LtuDXR, which is involved in terpenoid synthesis in Liriodendron. Our work lays the foundation for further exploration of the molecular mechanism by which terpenoid biosynthesis is regulated in Liriodendron.

Background The unique ‘mandarin jacket’ leaf shape is the most famous trait of Liriodendron chinense and this characteristic gives L. chinense aesthetic and landscaping value. However, the underlying regulatory mechanism of genes involved in the leaf development of L. chinense has remained unclear. Methods Based on transcriptome data of leaves at different developmental stages from L. chinense, we identified differentially expression genes (DEGs) functioning in leaf development. A candidate gene named LcCUC2-like (LcCUC2L) had high similarity in sequence with Arabidopsis thaliana CUC2, and used for further research. We isolated the full-length LcCUC2L gene and its promoter from L. chinense. Subsequently, we analyzed the function of the LcCUC2L gene and its promoter activity via transformation into A. thaliana. Results In this study, we found that the LcCUC2L and AtCUC2 are homologous in sequence but not homologous in function. Unlike the role of AtCUC2 in leaf serration and SAM formation, the LcCUC2L mainly regulates cotyledon development and rosette leaf number. Histochemical β-glucuronidase (GUS) staining revealed that LcCUC2L was expressed in the cotyledons of A. thaliana seedlings, indicating that the LcCUC2L may play a role in cotyledon development. Ectopic expression of LcCUC2L resulted in long, narrow cotyledons without petioles, abnormal lamina epidermis cells and defective vascular tissue in cotyledons, and these results were consistent with the LcCUC2L expression pattern. Further analysis showed that overexpression of LcCUC2L also induced numerous rosette leaves. Also, LcCUC2L and other related genes showed a severe response in L. chinense by introducing exogenous auxin stimulation, partly revealed that LcCUC2L affects the leaf development by regulating the auxin content. Conclusions These results suggest that LcCUC2L may play a critical role in leaf development and morphogenesis in L. chinense, and our findings provide insight into the molecular mechanisms of leaf development in L. chinense.