Molecular Mechanism of LFY and UFO Synergistically Regulating Flower Development

François Parcy's team published a research paper entitled "The F-box protein UFO controls flower development by redirecting the master transcription factor LEAFY to new cis-elements" on Nature Plants. This study found that an important role of UFO is to form a transcription complex with LFY on the new DNA motif LUBS to activate the expression of the main transcription factor AP3 in flower development; revealing the molecular mechanism of LFY and UFO synergistically regulating flower development.

Flowers consist of four organs (sepals, petals, stamens, and carpels) arranged in concentric rings. Flower development is key to the successful reproduction of angiosperms. Floral meristem (FM) patterning is regulated by ABCE floral homologous genes that determine the identity of individual floral organs. In Arabidopsis, LEAFY (LFY) is the main transcription factor that activates the ABCE gene. LFY uniformly activates the A gene APETALA1 (AP1) in early FM and activates the local expression of B and C genes together with other cofactors. LFY can regulate the C gene AGAMOUS (AG) together with WUSCHEL (WUS) to determine the development of stamens and carpels. In addition, the main function of LFY and UNUSUAL FLORAL ORGANS (UFO) is to co-activate the B gene APETALA3 (AP3), thereby controlling the development of petals and stamens. However, what is the molecular mechanism behind the synergy between LFY and UFO remains unclear.

Activation of floral development gene promoters by LFY and UFO was determined in Arabidopsis protoplasts by dual luciferase reporter (DLRA). This study found that UFOs with or without F-box deletion (UFO and UFOΔF-box) had very similar activation behaviors. Indicating that the role of the F-box domain is largely dispensable, LFY-UFO synergy should be regulated by a mechanism independent of ubiquitination.

Using electrophoretic mobility shift assays (EMSAs), this study found that UFO can form an active transcription complex with LFY and bind to a specific regulatory sequence (LUBS0) of the AP3 promoter. Further through genome-wide analysis, the researchers found that these novel regulatory elements contained either a single or two similar motifs compared to the LFY-binding sequence (LFYBS), termed mLUBS and dLUBS, respectively.

In addition to LUBS0, the study also found two other sequences that bind to the LFY-UFO complex, LUBS1 and LUBS2, and LUBS1 binds better to the LFY-UFO complex. By mutating the corresponding motifs, the researchers confirmed that LUBS1 and LUBS2 are required for the LFY-UFO complex to bind and induce AP3 gene expression. In addition, RBE, PI, and FD may also be bound by and regulated by the LFY-UFO complex.

In Arabidopsis, LFY has UFO-dependent and UFO-independent functions, respectively. The study found that the K249R mutation of LFY can specifically block the binding of LFY-UFO complex to its DNA motif, but does not affect the binding of LFY alone to its DNA motif. It indicated that Lys249 played a key role in the interaction between LFY-UFO and LUBS motif.

Through cryo-electron microscopy (cryo-EM), this study resolved the protein structure of the LFY-UFO-DNA complex at a resolution level of 4.27 Å. It also shows how UFO changes its transcription factor specificity, thereby mediating the synergistic effect of LFY and UFO, binding to specific DNA motifs in the form of complexes and regulating the expression of related genes. These results further demonstrate that UFOs perform their functions by directly interacting with LFY and DNA motifs.

Finally, through evolutionary analysis, the researchers found that the UFO-LFY complex also exists in non-angiosperm plants, and proposed that the complex may have a very early evolutionary origin, earlier than the formation of flowering plants.

In summary, this study discovered a new DNA motif LUBS that can be recognized by the UFO-LFY complex, identified the key site K249 for the binding of the UFO-LFY complex to DNA, and resolved the structural characteristics of the ASK1-UFO-LFY-DNA complex. It revealed that the F-box protein UFO can directly bind to the LUBS motif in the AP3 promoter and activate its expression by forming a complex with LFY, thereby specifically regulating the molecular mechanism of flower development.

Reference

Rieu, P., Turchi, L., Thévenon, E. et al. The F-box protein UFO controls flower development by redirecting the master transcription factor LEAFY to new cis-elements. Nat. Plants 9, 315–329 (2023). https://doi.org/10.1038/s41477-022-01336-2