Cloning of SMT Gene in Cynanchum auriculatum and Its Expression Analysis Under Abiotic Stresses
LIU Xin-Yue1, SU Xiao-Xue1, GUO Jia-Min1, ZANG Yue1, SUN Miao1,2, SHEN Min1,2, HONG Jian1,2, LI Xiang1,2, ZHANG Wen-Xuan1, LIU Fang-Fang1,2,*, KANG Yi-Jun1,2,*
1 School of Marine and Biological Engineering/Yancheng Bioengineering Research Center for 'Binhai Bai-shou-wu', Yancheng Teachers University, Yancheng 224007, China; 2 Jiangsu Key Laboratory for Bioresources of Saline Soils, Yancheng 224007, China
Abstract：Phytosterols play crucial roles in plant growth, development, and stress resistance, with C24-sterol methyltransferase (SMT) serving as a key rate-limiting enzyme in phytosterol biosynthesis pathway. Cynanchum auriculatum is rich in phytosterol-based bioactive components, to investigate the function of SMT in phytosterol biosynthesis and abiotic stress response in C. auriculatum, the genes encoding SMT were cloned based on transcriptome data, bioinformatic analysis was carried out, and the gene expression levels under abiotic stresses were detected by qRT-PCR. The results showed that 2 genes with ORF of 1 041 and 1 080 bp were cloned and named CaSMT1 (GenBank No. PV701631) and CaSMT2 (GenBank No. PV701632), respectively. CaSMT1 encoded a protein consisting of 346 amino acids, with a relative molecular weight of 38.65 kD and a theoretical isoelectric point of 6.33. CaSMT1 was a hydrophilic protein without signal peptide. CaSMT2 encoded a protein consisting of 359 amino acids, with a relative molecular weight of 40.16 kD and a theoretical isoelectric point of 6.42. CaSMT2 protein contained transmembrane domain and was localized to the plasma membrane. Both CaSMT proteins contained canonical sterol C-24 methyltransferase C-terminal domains, with secondary structures predominantly composed of alpha helix and random coil. Phylogenetic analysis revealed that CaSMT1 and CaSMT2 clustered into distinct clades, both exhibiting the closest phylogenetic relationship with their corresponding SMT in Calotropis procera. qRT-PCR analysis revealed distinct tissue-specific expression patterns for both genes, with significantly higher expression levels in roots and leaves compared with stems. Furthermore, steroidal glycoside accumulation exhibited concordant variation patterns with gene expression levels across all examined tissues. Under various abiotic stresses including salt, drought, heat, and cold, CaSMT1 exhibited significant upregulation across all tested conditions. In contrast, CaSMT2 showed distinct stress-responsive patterns: Marked upregulation under heat and cold stress, downregulation under drought stress, and no significant response to salinity. This differential regulation demonstrated the involvement of CaSMT paralogs in adaptive responses of C. auriculatum to diverse abiotic stresses. This study provides theoretical insights for improving stress resistance and enhancing the content of active component in medicinal plants.
刘鑫悦, 苏晓雪, 郭佳敏, 臧悦, 孙淼, 沈敏, 洪键, 李祥, 张文轩, 刘方方, 康贻军. 耳叶牛皮消SMT基因克隆及其在非生物胁迫下的表达分析[J]. 农业生物技术学报, 2025, 33(10): 2149-2163.
LIU Xin-Yue, SU Xiao-Xue, GUO Jia-Min, ZANG Yue, SUN Miao, SHEN Min, HONG Jian, LI Xiang, ZHANG Wen-Xuan, LIU Fang-Fang, KANG Yi-Jun. Cloning of SMT Gene in Cynanchum auriculatum and Its Expression Analysis Under Abiotic Stresses. 农业生物技术学报, 2025, 33(10): 2149-2163.
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