De novo organogenesis, which gives rise to adventitious roots and shoots, is a type of plant regeneration for survival after wounding. In Arabidopsis (Arabidopsis thaliana), two main cell fate transition steps are required to establish the root primordium during de novo root organogenesis from leaf explants. The first step from regeneration-competent cells to root founder cells involves activation of WUSCHEL-RELATED HOMEOBOX11 (WOX11) and WOX12 (WOX11/12) expression by auxin. However, the molecular mechanism controlling the second step of fate transition from root founder cells to root primordium is poorly understood. In this study, we show that the expression levels of WOX11/12 decrease while those of WOX5 and 7 (WOX5/7) increase during the transition from root founder cells to the root primordium. WOX11/12 function genetically upstream of WOX5/7, and the WOX11/12 proteins directly bind to the promoters of WOX5/7 to activate their transcription. Mutations in WOX5/7 result in defective primordium formation. Overall, our data indicate that the expression switch from WOX11/12 to WOX5/7 is critical for initiation of the root primordium during de novo root organogenesis.