To investigate the possible mechanism(s) by which dietary manganese (Mn) levels and sources modulate the expression of the manganese-containing superoxide dismutase (MnSOD) gene at both the transcriptional and translational levels, we used 432 8-d-old male broiler chicks in a 1 plus 4 × 2 design. Chickens were given either a diet without Mn supplementation [control (C)] or diets supplemented with 100 (optimal) or 200 (high) mg Mn/kg diet from inorganic Mn sulfate (I) or 3 organic complexes of Mn and amino acids with weak (W), moderate (M), or strong (S) chelation strength up to 21 d of age. Compared with C chicks, chicks fed Mn-supplemented diets had higher (P < 0.01) Mn concentrations, specificity protein 1 (Sp1) DNA-binding activities, MnSOD mRNA levels, MnSOD mRNA-binding protein (MnSOD-BP) RNA-binding activities, MnSOD protein concentrations, and MnSOD activities within heart tissue, but lower (P < 0.01) heart activating protein-2 (AP-2) DNA-binding activities. Chicks fed M diets had higher (P < 0.05) heart Mn concentrations, MnSOD mRNA levels, and MnSOD-BP RNA-binding activities compared with those fed the I and W diets and lower (P < 0.01) AP-2 DNA-binding activities than those fed other treatment diets. These results suggest that dietary Mn could modulate the expression of the MnSOD gene in broilers by altering Sp1 and AP-2 DNA-binding activities at the transcriptional level and enhancing MnSOD-BP RNA-binding activity at the translational level. Additionally, an organic Mn source with moderate chelation strength could be more effective than other Mn sources in activating MnSOD gene expression at both the transcriptional and translational levels.