The core plant circadian oscillator is composed of multiple interlocked transcriptional-translational feedback loops, which synchronize endogenous diel physiological rhythms to the cyclic changes of environmental cues. PSEUDO-RESPONSE REGULATORS (PRRs) have been identified as negative components in the circadian clock, though their underlying molecular mechanisms remain largely unknown. Here we found that a subfamily of zinc finger transcription factors, B-box (BBX)-containing proteins, have a critical role in fine-tuning circadian rhythm. We demonstrated that overexpressing Arabidopsis thaliana BBX19 and BBX18 significantly lengthened the circadian period, while the null mutation of BBX19 accelerated the circadian speed. Moreover, BBX19 and BBX18, which are expressed during the day, physically interacted with PRR9, PRR7, and PRR5 in the nucleus in precise temporal ordering from dawn to dusk, consistent with the respective protein accumulation pattern of PRRs. Our transcriptomic and genetic analysis indicated that BBX19 and PRR9, PRR7, and PRR5 cooperatively inhibited the expression of morning-phased clock genes. PRR proteins affected BBX19 recruitment to the CCA1, LHY, and RVE8 promoters. Collectively, our findings show that BBX19 interacts with PRRs to orchestrate circadian rhythms, and suggest the indispensable role of transcriptional regulators in fine-tuning the circadian clock.
Four Holstein cows fitted with ruminal and T-type duodenal cannulas were utilized in a 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments. The TMR contained 25% alfalfa haylage, 25% corn silage, and 50% concentrate and provided either 16.4 or 19.6% CP, with ruminal degradability calculated to be 30 or 45%. Intakes of DM, OM, ADF, NDF, and N were not altered by either amount or degradability of CP. Intake and ruminal and postruminal digestibility of starch were greater when cows were fed diets high in undegradable CP but was not altered by amount of CP. Apparent total tract digestibilities for DM, OM, starch, ADF, and NDF were similar among treatments. Apparent total tract digestibility of N was 4.7 percentage units greater for diets low in ruminally degradable CP. Apparent digestibility of OM, ADF, and NDF and true digestibility of OM in the rumen were not altered by amount of CP or undegradable CP. Increasing the CP content of the diet and the proportion of undegradable CP in the diet increased NAN flow to the duodenum. Except for Met, flows of all AA to the duodenum were increased when CP was increased. Flow of Met to the duodenum was not altered by undegradable CP content of the diet. Production of milk, 4% FCM, and milk CP was not altered by amount of CP or undegradable CP. Milk fat content and yield were increased when diets high in undegradable CP were fed. Results suggest that all diets supplied adequate amounts of AA for these cows or that Met was deficient for all cows.
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