DWF4 encodes a rate-limiting mono-oxygenase that mediates 22alpha-hydroxylation reactions in the BR biosynthetic pathway and it is the target gene in the BR feedback loop. Knockout of DWF4 results in a dwarfed phenotype and other severe defects in Arabidopsis. Here we report on the isolation of the ZmDWF4 gene in maize. Sequence analysis revealed that the open reading frame of ZmDWF4 was 1,518 bp, which encodes a protein composed of 505 amino acid residues with a calculated molecular mass of 57.6 kD and a predicated isoelectric point (pI) of 9.54. Phylogenetic analysis indicated that ZmDWF4 was very close to the Arabidopsis DWF4. In young maize seedlings, the expression of ZmDWF4 in shoots was much higher than that in roots. The highest expression of ZmDWF4 was observed in husk leaves and the lowest in silks during flowering stage. The expression of ZmDWF4 in maize was significantly down regulated by exogenous brassinolide. A heterogeneous complementary experiment demonstrated that the defects of three Arabidopsis DWF4 mutants could be rescued by constitutive expression of ZmDWF4, with leaf expandability, inflorescence stem heights and fertile capabilities all restored to normal levels. Increases in seed and branch number as well as the height of florescence stem were observed in the over-expressed transformants. These findings suggest that ZmDWF4 may be an ortholog gene of Arabidopsis DWF4 and responsible for BR biosynthesis in maize.