Objective Based on transcriptome data from roots of Panax notoginseng under arsenic (As) stress, the tonoplast intrinsic protein gene PnTIP1;3 was cloned. Its functional characteristics in As transport were investigated through bioinformatic analysis and heterologous expression in yeast. Methods Bioinformatic tools were used to analyze the sequence features and promoter cis-acting elements of PnTIP1;3. A yeast heterologous expression system was constructed, and the role of PnTIP1;3 in cellular As transport was systematically evaluated through phenotypic assays, intracellular As content and subcellular distribution measurement by ICP-MS, and analysis of relevant antioxidant indicators. Results The tonoplast intrinsic protein gene PnTIP1;3 was cloned, with an open reading frame of 750 bp encoding 250 amino acids. Phylogenetic analysis indicated that it belongs to the TIP1 subfamily. Promoter analysis revealed the presence of multiple cis-acting elements related to hormones, growth and development, and stress responses. As tolerance assays showed that the half-maximal effective concentration (EC₅₀) of yeast transformants heterologously expressing PnTIP1;3 (TPnTIP1;3) was 5.36 mmol/L, higher than that of the empty vector control (5.03 mmol/L). As treatment inhibited the growth of both strains, but the empty vector control was more severely affected. ICP-MS analysis indicated that total As accumulation was significantly higher in TPnTIP1;3 than in the control, and the subcellular distribution of As followed the pattern: cell wall > vacuole > cytoplasm. Under As stress, reactive oxygen species (ROS) levels were significantly decreased (P < 0.001) in TPnTIP1;3, while the activities of superoxide dismutase (SOD) and catalase (CAT), as well as the contents of glutathione (GSH) and metallothionein (MT), were significantly increased (P < 0.05). Correlation analysis further demonstrated significant positive relationships between these antioxidant indicators and both total As content and cell wall As concentration. Conclusion The P. notoginseng tonoplast intrinsic protein gene PnTIP1;3, a member of the TIP1 subfamily, enhances yeast tolerance to As stress by promoting As accumulation and vacuolar sequestration, and through activation of the host antioxidant system.

Panax notoginseng(Burk) F.H.Chen  /  aquaporin  /  arsenic  /  bioinformatics analysis  /  heterologous expression  /  antioxidant system