Mineral nutrient deficiencies and metal ion toxicities coexist on acid soils. Phosphorus (P) deficiency in plants is generally accompanied with significant levels of leaf manganese (Mn) accumulation. However, the molecular regulatory mechanisms underpinning remain unclear. The present study found that P-deficient soybean plants accumulated more Mn compared to P-sufficient ones on acid soils in both field and greenhouse experiments. Meanwhile, both P deficiency and Mn toxicity enhanced the expression of GmSTOP1-3. Over-expressing GmSTOP1-3 enhanced Mn accumulation in transgenic soybean hairy roots, but RNA-interference did not show obvious differences. Moreover, transgenic soybeans with GmSTOP1-3-overexpression showed enhanced root citrate exudation and augmented Mn accumulation. RNA-sequence identified four downstream genes of GmSTOP1-3, including multidrug and toxic compound extrusion (GmMATE2/13) and metal transporter genes (GmZIP6/GmIREG3), which encode plasma membrane proteins. GmSTOP1-3 activated the transcription of these four genes by directly binding to their promoter regions. In addition, both GmZIP6 and GmIREG3 functioned in Mn uptake as manifested by the higher Mn concentration and decreased growth of soybean hairy roots with their overexpression. Taken together, it is suggested that upregulation of GmSTOP1-3 by low P stress on acid soils activates transcripts of GmMATE2/13 and GmZIP6/GmIREG3, which consequently result in enhanced Mn accumulation in soybean.