Intercellular protein movement plays a critical role in animal and plant development. SHORTROOT (SHR) is a moving transcription factor essential for endodermis specification in the Arabidopsis root. Unlike diffusible animal morphogens, which form a gradient across multiple cell layers, SHR movement is limited to essentially one cell layer. However, the molecular mechanism is unknown. We show that SCARECROW (SCR) blocks SHR movement by sequestering it into the nucleus through protein-protein interaction and a safeguard mechanism that relies on a SHR/SCR-dependent positive feedback loop for SCR transcription. Our studies with SHR and SCR homologs from rice suggest that this mechanism is evolutionarily conserved, providing a plausible explanation why nearly all plants have a single layer of endodermis.

The development of multicellular organisms relies on the coordinated control of cell divisions leading to proper patterning and growth [1][2][3] . The molecular mechanisms underlying pattern formation, particularly the regulation of formative cell divisions, remain poorly understood. In Arabidopsis, formative divisions generating the root ground tissue are controlled by SHORTROOT (SHR) and SCARECROW (SCR) 4-6. Here we show, using cell-type-specific transcriptional effects of SHR and SCR combined with data from chromatin immunoprecipitation-based microarray experiments, that SHR regulates the spatiotemporal activation of specific genes involved in cell division. Coincident with the onset of a specific formative division, SHR and SCR directly activate a D-type cyclin; furthermore, altering the expression of this cyclin resulted in formative division defects. Our results indicate that proper pattern formation is achieved through transcriptional regulation of specific cellcycle genes in a cell-type-and developmental-stage-specific context. Taken together, we provide evidence for a direct link between developmental regulators, specific components of the cell-cycle machinery and organ patterning.Growth and patterning are key processes that govern the development of multicellular organisms. In some cases, like early Drosophila embryogenesis 7 , these are independent. However, in many animals and plants, proper development frequently relies on tight coordination of growth and patterning. Disruption of this coordination can lead to unchecked cell growth, resulting in tumorigenesis or misshapen organs 8 . Although the molecular mechanisms involved in pattern formation 9-11 and in cell-cycle control [12][13][14][15] 5,6,23,24,25 . To gain insight into the role of the SHR/SCR network in controlling formative cell divisions, we expressed an inducible version of either SHR or SCR in its respective mutant background and characterized the timing of formative divisions after induction.Before induction, SHR-and SCR-inducible plants had a single mutant ground tissue layer 4,5 (Supplementary Fig. 1). After SHR induction, SCR expression was observed within 3 h, indicating that SHR rapidly activates its targets ( Supplementary Fig. 1). The first periclinal (parallel to the direction of growth) division in the mutant ground tissue layer occurred 6 h after SHR induction (Fig. 1a , Supplementary Fig. 1 and Supplementary Movie 1) and earlier after SCR induction (Fig. 1b). Two layers of ground tissue with SCR expression in the quiescent centre and endodermis (Supplementary Fig. 1), along with a nearly complete Casparian band 5 , were detected 24 h after SHR induction ( Supplementary Fig. 2). This underlines the combinatorial role of SHR and SCR in regulating formative cell divisions and also indicates that the two inducible systems have slightly different kinetics.To understand the dynamics of the SHR/SCR regulatory network, we sorted ground tissue cells at several time points after SHR and SCR induction and performed microarray a...