N⁶-methyladenosine (m⁶A) is becoming one of the most important RNA modifications in plant growth and development, including defense, cell differentiation, and secondary metabolism. YT521-B homology (YTH) domain-containing RNA-binding proteins, identified as m⁶A readers in epitranscriptomics, could affect the fate of m⁶A-containing RNA by recognizing and binding the m⁶A site. Therefore, the identification and study of the YTH gene family in Liriodendron chinense (L. chinense) can provide a molecular basis for the study of the role of m⁶A in L. chinense, but studies on the YTH gene in L. chinense have not been reported. We identified nine putative YTH gene models in the L. chinense genome, which can be divided into DF subgroups and DC subgroups. Domain sequence analysis showed that the LcYTH protein had high sequence conservation. A LcYTH aromatic cage bag is composed of tryptophan and tryptophan (WWW). PrLDs were found in the protein results of YTH, suggesting that these genes may be involved in the process of liquid–liquid phase separation. LcYTH genes have different tissue expression patterns, but the expression of LcYTHDF2 is absolutely dominant in all tissues. In addition, the expression of the LcYTH genes is changed in response to ABA and MeJA. In this study, We identified and analyzed the expression pattern of LcYTH genes. Our results laid a foundation for further study of the function of the LcYTH gene and further genetic and functional analyses of m⁶A RNA modification in forest trees.

Stress-associated proteins (SAPs), belonging to the A20/AN1 zinc finger protein family, are key regulators in plant stress responses. Despite their importance, studies on the SAP gene family in Pinus massoniana are still relatively scarce. This study aimed to systematically identify and characterize SAP genes in P. massoniana and to explore their potential roles in stress response mechanisms. A total of 17 PmSAP genes were identified from P. massoniana. Phylogenetic analysis revealed that these genes group into five distinct clades, and 10 conserved motifs were identified. Using transcriptome data and qRT-PCR, we analyzed their expression patterns and employed yeast systems to validate their transcriptional activities. The responses of PmSAP gene family members to different stress treatments showed significant differences. For example, PmSAP8 and PmSAP12 responded strongly to ABA, MeJA, and H₂O₂ treatments, while PmSAP3 and PmSAP5 showed significant upregulation under ETH and NaCl stress. Yeast experiments indicated that PmSAP6/8/12 were transcriptional activators, and PmSAP3 and PmSAP5 were transcriptional suppressors. The identification and preliminary analysis of PmSAP genes provided a theoretical foundation for understanding stress resistance mechanisms in P. massoniana.

Terpenoids, abundant and structurally diverse secondary metabolites in plants, especially in conifer species, play crucial roles in the plant defense mechanism and plant growth and development. In Pinus massoniana, terpenoids’ biosynthesis relies on both the mevalonate (MVA) pathway and the 2-methyl-D-erythritol-4-phosphate (MEP) pathway, with 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate synthase (HDS) catalyzing the sixth step of the MEP pathway. In this study, we cloned and conducted bioinformatics analysis of the PmHDS gene from P. massoniana. The results showed that PmHDS shares homology with HDS proteins from other species. Analysis of tissue expression patterns indicated that PmHDS exhibits the highest expression level in xylem tissue, followed by stems, with significantly lowest expression in the apical meristem. Treatment with NaCl, abscisic acid (ABA), ethylene (ETH), methyl jasmonate (MeJA), and salicylic acid (SA) upregulated the expression of PmHDS. Furthermore, we successfully cloned the PmHDS promoter (about 2220 bp) and integrated it into a GUS reporter vector, which resulted in GUS activity being observed in various tissues of Arabidopsis thaliana. Overexpression of the PmHDS gene in A. thaliana significantly increased the content of carotenoids, chlorophylls a and b, and related enzyme activities, as well as the levels of terpenoid derivatives such as cytokinin (CTK), gibberellic acid (GA), and ABA, thereby enhancing the resistance to those abiotic stresses. These findings suggest that PmHDS plays an important role in the terpenoid synthesis pathway. This study provides a theoretical basis for understanding the biosynthesis of terpenoids and lays a foundation for future research on the regulation of terpene synthesis and resistance in molecular breeding.

Transcription factors refer to types of proteins that perform significant functions in the process of gene expression regulation. The ethylene insensitive 3/ethylene insensitive 3-like (EIN3/EIL) family, functioning as significant transcription factors regulating ethylene, plays a critical role in the growth and development of plants and participates in the plant’s response to diverse environmental stresses. Pinus massoniana is an excellent native tree with high economic and ecological value. However, the study of EIN3/EIL genes in gymnosperms, for instance, P. massoniana, is still relatively limited. In this research, four putative EIN3/EIL genes were identified in the transcriptome of P. massoniana. Bioinformatics analysis showed that PmEIL genes contain a highly conserved EIN3 domain and other structural features of acidic, proline-rich and glutamine-rich sites. The molecular evolution tree analysis demonstrated that the EIN3/EIL family was partitioned into three categories (A, B, and C), and the number, type, and distribution of conserved motifs grouped in one category were similar. The results of qRT-PCR indicated that the expression levels of PmEIL genes were markedly elevated in needles compared to other tissues. Through the analysis of expression patterns of the PmEIL genes under various stress treatments, it was found that the PmEIL genes could participate in plant hormone stimulation induction, osmosis, drought and other response processes. In addition, PmEIL is a nuclear localization protein. PmEIL1, PmEIL3, and PmEIL4 are transcriptional activators, while PmEIL2 is a transcriptional suppressor. This research provides a basis for further elucidating the function of EIN3/EIL transcription factors in growth, development and stress response of P. massoniana.