Genome-Wide Identification and Functional Analysis of Salvia miltiorrhiza MicroRNAs Reveal the Negative Regulatory Role of Smi-miR159a in Phenolic Acid Biosynthesis.

MicroRNAs (miRNAs) are a group of endogenous small non-coding RNAs in plants. They play critical functions in various biological processes during plant growth and development. Salvia miltiorrhiza is a well-known traditional Chinese medicinal plant with significant medicinal, economic, and academic values. In order to elucidate the role of miRNAs in S. miltiorrhiza , six small RNA libraries from mature roots, young roots, stems, mature leaves, young leaves and flowers of S. miltiorrhiza and one degradome library from mixed tissues were constructed. A total of 184 miRNA precursors, generating 137 known and 49 novel miRNAs, were genome-widely identified. The identified miRNAs were predicted to play diversified regulatory roles in plants through regulating 891 genes. qRT-PCR and 5' RLM-RACE assays validated the negative regulatory role of smi-miR159a in SmMYB62 , SmMYB78 , and SmMYB80 . To elucidate the function of smi-miR159a in bioactive compound biosynthesis, smi-miR159a transgenic hairy roots were generated and analyzed. The results showed that overexpression of smi-miR159a caused a significant decrease in rosmarinic acid and salvianolic acid B contents. qRT-PCR analysis showed that the targets of smi-miR159a, including SmMYB62 , SmMYB78, and SmMYB80 , were significantly down-regulated, accompanied by the down-regulation of SmPAL1 , SmC4H1 , Sm4CL1 , SmTAT1 , SmTAT3 , SmHPPR1 , SmRAS, and SmCYP98A14 genes involved in phenolic acid biosynthesis. It suggests that smi-miR159a is a significant negative regulator of phenolic acid biosynthesis in S. miltiorrhiza .