The WOX9-WUS modules are indispensable for the maintenance of stem cell homeostasis in Arabidopsis thaliana.

The dynamic balance between the self-renewal and differentiation of stem cells in plants is precisely regulated by a series of developmental regulated genes that exhibit spatiotemporal-specific expression patterns. Several studies have demonstrated that the WOX family transcription factors play critical roles in maintaining the identity of stem cells in Arabidopsis thaliana. In this study, we obtained amiR-WOX9 transgenic plants, which displayed terminating prematurely of shoot apical meristem (SAM) development, along with alterations in inflorescence meristem and flower development. The phenotype of amiR-WOX9 plants exhibited similarities to that of wus-101 mutant, characterized by a stop-and-go growth pattern. It was also found that the expression of WUS in amiR-WOX9 lines was decreased significantly, while in UBQ10::WOX9-GFP transgenic plants, the WUS expression was increased significantly despite no substantial alteration in meristem size compared to Col. Therefore, these data substantiated the indispensable role of WOX9 in maintaining the proper expression of WUS. Further investigations unveiled the direct binding of WOX9 to the WUS promoter via the TAAT motif, thereby activating its expression. It was also found that WUS recognized identical the same TAAT motif cis-elements in its own promoter, thereby repress self-expression. Next, we successfully identified a physical interaction between WOX9 and WUS, and verified that it was harmful to the expression of WUS. Finally, our experimental findings demonstrate that WOX9 was responsible for the direct activating of WUS, which however was interfered by the ways of WUS binding its own promoter and the interaction of WUS and WOX9, thereby ensuring the appropriate expression pattern of WUS and then the stem cell stability. This study contributes to an enhanced comprehension of the regulatory network of the WOX9-WUS module in maintaining the equilibrium of the SAM.