BdDOF24 interacting with BdGAMYB regulates the BdCathB gene upon germination. During barley seed germination, hydrolytic enzymes (α-amylases, proteases, etc.) synthesized in the aleurone layer in response to gibberellins (GA), catalyse the mobilization of storage reserves accumulated in the endosperm during seed maturation. In Brachypodium distachyon, the BdCathB gene that encodes a Cathepsin B-like thiol-protease, orthologous to the wheat Al21 and barley HvCathB, is highly induced in germinating seeds and its expression is regulated by transcription factors (TFs) encoded by genes BdGamyb and BdDof24, orthologous to the barley HvGamyb and BPBF-HvDof24, respectively. Transcripts of both TF genes increase during germination and treatments with abscisic acid (ABA) or paclobutrazol (PAC, an inhibitor of GA biosynthesis) decrease mRNA expression of BdGamyb but do not affect that of BdDof24. Besides, proteins BdDOF24 and BdGAMYB interact in yeast-2 hybrid systems and in plant nuclei, and in transient expression assays in aleurone layers BdDOF24 is a transcriptional repressor and BdGAMYB is an activator of the BdCathB promoter, as occurs with the putative orthologous in barley BPBF-HvDOF24 and HvGAMYB. However, when both TFs are co-bombarded, BdDOF24 enhances the activation driven by BdGAMYB while BPBF-HvDOF24 strongly decreases the HvGAMYB-mediated activation of the BdCathB promoter. The different results obtained when BdDOF24 and BPBF-HvDOF24 interact with BdGAMYB and HvGAMYB are discussed.