[Cloning, expression pattern, and functional verification of phosphate transporter protein SmPht5;1 in Salvia...

Vacuolar phosphate transporters have recently been identified as key regulators of intracellular phosphate status in plant cells. However, related studies have not yet been reported on them in Salvia miltiorrhiza. In response, specific amplification primers were designed according to the open reading frame sequence of SmPht5;1 obtained from the S. miltiorrhiza genome, and the full-length SmPht5;1 gene was cloned. Bioinformatics analysis was conducted to characterize the physicochemical properties, subcellular localization, and conserved domains of its encoded protein. The secondary and tertiary structures were predicted, and a phylogenetic tree was constructed. Furthermore, transcriptome data were analyzed to investigate the expression patterns of this gene in different tissues of S. miltiorrhiza and under varying phosphate levels. A yeast complementation assay was employed to validate its phosphate transport function. The results showed that the SmPht5;1 gene was 2 085 bp in length, encoding 694 amino acids. The encoded phosphate transporter had a relative molecular mass of 77 412.45 and was classified as a hydrophobic protein. Its secondary structure was predominantly composed of α-helices and random coils. The SmPht5;1 protein sequence contained complete SPX and MFS domains, consistent with the structural characteristics of vacuolar phosphate transporters, and exhibited high sequence similarity with PHT5 subfamily proteins from other plants, particularly showing 72.57% similarity with AtPht5;1. Phylogenetic analysis indicated that SmPht5;1 likely functioned in transporting phosphate into the vacuole. Transcriptome data analysis revealed that SmPht5;1 was expressed in all tissues of S. miltiorrhiza, with the highest expression level in leaves. Under low-phosphate stress, the expression of SmPht5;1 was downregulated in the aerial parts. Yeast functional validation results demonstrated that SmPht5;1 can complement the function of yeast mutant strains, confirming its phosphate transport capability. This study provides a theoretical foundation for improving phosphate utilization efficiency and redistribution in cultivated S. miltiorrhiza, holding significant practical importance for achieving high yield and quality in S. miltiorrhiza production.