Unveiling previously undescribed circadian clock regulators and action mechanisms via TurboID-based profiling of the LWD1 interactome in <i>Arabidopsis thaliana</i>.

Abstract

The synchronization of the internal circadian clock with the external 24-h diel cycle is critical for plant growth and fitness. LIGHT-REGULATED WD proteins (LWDs) are essential for sustaining a precise and robust circadian clock in <i>Arabidopsis</i> [Y. Wang <i>et al.</i>, <i>Plant Cell</i> <b>23</b>, 486-498 (2011); J. F. Wu <i>et al.</i>, <i>Nat Commun.</i> <b>7</b>, 13181 (2016)]. However, the action mechanisms by which LWDs orchestrate the expression of clock genes remain largely elusive. We systematically identified 219 LWD1-interacting protein partners at three Zeitgeber times (ZTs) using a TurboID-mediated proximity labeling approach. Selected interacting proteins were experimentally verified as new regulators of the <i>Arabidopsis</i> circadian clock. Our data also support that LWD1-PSEUDO RESPONSE REGULATORS (PRRs)-TOPLESS (TPL) complex represses the morning gene <i>CIRCADIAN CLOCK ASSOCIATED</i> (<i>CCA1</i>), shaping the expression trough of <i>CCA1</i> in the late afternoon/early evening under diel cycles. Our results revealed a new corepressor role of LWD1, working in conjunction with epigenetic regulators and transcription factors to underpin the precise operation of the circadian clock.