Phase separation, the formation of biomacromolecular structures without membranes inside cells, plays a crucial role in organismal development and signal transduction. The plant photoperiod regulatory factor CONSTANS (CO) protein can interact with nuclear transcription factors NF-YB2 and NF-YC9 to co-activate the expression of the key flowering gene FLOWERING LOCUS T (FT), thereby promoting the floral transition in plants. However, the extent to which phase separation contributes to this mechanism and the precise molecular underpinnings are yet to be elucidated.
Recently, researchers from the South China Botanical Garden of the Chinese Academy of Sciences and Nanyang Technological University in Singapore, led by Professors HOU Xingliang and Miao Yansong, respectively, have revealed the specific regulatory mechanism of phase separation in plant flowering through collaborative research. The study was published in The EMBO Journal. Dr. HUANG Xiang, a postdoctoral fellow at the South China Botanical Garden, and Dr. MA Zhiming, a postdoctoral fellow at Nanyang Technological University, were the co-first authors of the paper, while Professor Hou and Professor Miao were the co-corresponding authors.
The study found that CO, NF-YB2, and NF-YC9 proteins can form phase-separated complexes in plants. To further analyze the specific formation mechanism and regulatory function, researchers utilized protoplasts, in vitro experimental systems, and transgenic plants to detect the regulation of these transcription factors on FT gene expression. Results showed that in response to light signals, CO proteins form functional percolation clusters from a diffuse distribution in a B-box-motif-dependent manner. Multivalent co-assembly with NF-YC9 and NF-YB2 is necessary to maintain proper CO assembly and transcriptional activity, preventing the formation of CO inhibitory slow-diffusing condensates that would impede FT activation.
To elucidate how NF-Y factors maintain the fluidity of CO/NF-YB2/NF-YC9 condensates for transcriptional activation of the flowering genes, researchers discovered that the intrinsically disordered region (IDR) of NF-YC9 contains a polyglutamine (polyQ) motif, fine-tunes the functional properties of CO/NF-YB/NF-YC condensates. Specific recognition of the FT promoter and polyelectrolyte partitioning enable the fluidic functional properties of CO/NF-YB/NF-YC/FT condensates, thereby allowing transcriptional activation of FT.
This study reveals the critical role of phase separation in the floral transition, clarifying how plants precisely control the flowering process by regulating the state of transcription factor condensates (Figure 1). It provides new insights into the photoperiodic flowering pathway mechanism in plants.
The research was funded by the Guangzhou Science and Technology Program and the Guangdong Provincial Key Research and Development Program. The paper can be accessed at: https://www.embopress.org/doi/full/10.1038/s44318-024-00293-0.
Figure 1. A model of fine-tuned CO condensation in regulating FT expression and flowering.(Image by HOU et al)
The condensation and function of CO are driven by multivalent interactions. Coassembly with NF-YC9, NF-YB2, and FT is essential for the formation of percolation clusters of CO condensates, which maintain functional fluidic properties required for activating FT transcription and triggering flowering transition. However, in the absence of NF-YC, CO proteins lead to over-assembly via strong self-interaction into slow-fluidic condensates, resulting in the improper partition of flexible NF-YB proteins and leading to the formation of malfunctional gelated condensates.