Abstract: SmMYB52, a transcription factor belonging to the R2R3-MYB subfamily of Salvia miltiorrhiza Bunge, can promote the accumulation of salvianolic acid B and inhibit root growth and development. Protein truncation analysis and transactivation assays were conducted to characterize regulatory activity of SmMYB52. Interacting proteins were identified through screening of a yeast two-hybrid cDNA library, and candidate interactions were subsequently validated using luciferase complementation imaging and bimolecular fluorescence complementation assays. Strong transcriptional self-activation activity was detected in the C-terminal region of SmMYB52. Library screening with SmMYB52-N-BD as a bait plasmid identified 15 candidate proteins capable of interacting with SmMYB52, including transcription factors SmTCP31 and SmTCP23 as well as lectin SMLⅡ. Subsequent validation experiments demonstrated a direct protein-protein interaction between SmMYB52 and SmTCP31. These findings provide a foundation for further investigation of the molecular mechanisms associated with SmMYB52 in S. miltiorrhiza.