B-Box proteins BBX28 and BBX29 interplay with PSEUDO-RESPONSE REGULATORS to fine-tune circadian clock in Arabidopsis

As an endogenous time-keeping mechanism, the circadian clock benefits plant fitness and adaptation to the rhythmically changed diel environments. The key components within the core oscillator of plant circadian clock have been extensively characterised, however, the fine-tuning circadian regulators are still less identified. Here, we demonstrated that BBX28 and BBX29, the two B-Box V subfamily members lacking DNA-binding motifs, are involved in the regulation of Arabidopsis circadian clock. Over-expressing either BBX28 or BBX29 significantly lengthened circadian period, whereas loss-of-function of BBX28 rather than BBX29 displayed a modestly long period in free-running condition. Mechanistically, BBX28 and BBX29 interacted with core clock components PRR5, PRR7 and PRR9 in nucleus to augment their transcriptional repressive activities. RNA-sequencing analysis further revealed that BBX28 and BBX29 shared 686 common differentially expressed genes (DEGs) including a subset of known direct transcriptional targets of PRR proteins within core oscillator, including CCA1, LHY, LNKs and RVE8 etc. Intriguingly, PRR proteins can feedback repress BBX28 and BBX29 transcription by associating with their promoters. Together, our findings unmasked an exquisite mechanism in which BBX28 and BBX29 interplay with PRR proteins to fine-tune the circadian pace.