Arabidopsis F-BOX STRESS INDUCED 4 is Required to Repress Excessive Divisions in Stomatal Development

During the terminal stage of stomatal development,the R2 R3-MYB transcription factors FOUR LIPS(FLP/MYB124) and MYB88 limit guard mother cell division by repressing the transcript levels of multiple cell-cycle genes. In Arabidopsis thaliana possessing the weak allele flp-1, an extra guard mother cell division results in two stomata having direct contact.Here, we identified an ethylmethane sulfonatemutagenized mutant, flp-1 xs01 c, which exhibited more severe defects than flp-1 alone, producing gianttumor-like cell clusters. XS01 C, encoding F-BOX STRESS-INDUCED 4(FBS4), is preferentially expressed in epidermal stomatal precursor cells.Overexpressing FBS4 rescued the defective stomatal phenotypes of flp-1 xs01 c and flp-1 mutants. The deletion or substitution of a conserved residue(Proline166) within the F-box domain of FBS4 abolished or reduced, respectively, its interaction with Arabidopsis Skp1-Like1(ASK1), the core subunit of the Skp1/Cullin/F-box E3 ubiquitin ligase complex. Furthermore, the FBS4 protein physically interacted with CYCA2;3 and induced its degradation through the ubiquitin-26 S proteasome pathway. Thus, in addition to the known transcriptional pathway, the terminal symmetric division in stomatal development is ensured at the post-translational level, such as through the ubiquitination of target proteins recognized by the stomatal lineage F-box protein FBS4.