Molecular mechanisms of Evening Complex activity in Arabidopsis

The Evening Complex (EC), composed of the DNA-binding protein LUX ARRHYTHMO (LUX) and two additional proteins, EARLY FLOWERING 3 (ELF3) and ELF4, is a transcriptional repressor complex and a core component of the plant circadian clock. In addition to maintaining oscillations in clock gene expression, the EC also participates in temperature and light entrainment and regulates important clock output genes such as PHYTOCHROME INTERACTING FACTOR 4 (PIF4), a key transcription factor involved in temperature dependent plant growth. These properties make the EC an attractive target for altering plant development through targeted mutations to the complex. However, the molecular basis for EC function was not known. Here we show that binding of the EC requires all three proteins and that ELF3 decreases the ability of LUX to bind DNA whereas the presence of ELF4 restores interaction with DNA. To be able to manipulate this complex, we solved the structure of the DNA-binding domain of LUX bound to DNA. Using structurebased design, a LUX variant was constructed that showed decreased in vitro binding affinity but retained specificity for its cognate sequences. This designed LUX allele modulates hypocotyl elongation and flowering. These results demonstrate that modifying the DNAbinding affinity of LUX can be used to titrate the repressive activity of the entire EC, tuning growth and development in a predictable manner.