GSE134333
Circadian rhythms in the absence of the clock gene Bmal1
Circadian clocks exist in almost all levels of living organisms and play elementary roles in the persistence of regular physiological and behavioural processes. Canonical transcription/translation feedback loop models portray BMAL1 (ARNTL) as one of the principal drivers of circadian periodicity in mammalian systems. In this integrated multi-omics study, we demonstrate, for the first time, 24 hr circadian oscillations in the expression levels of several transcripts and proteins in dexamethasone-synchronized Bmal1-/- mouse fibroblast cells and liver tissue slices. Intriguingly, daily oscillations were also observed in phosphoproteome profiles in the absence of this core clock gene. Our findings reveal that Bmal1 knockout radically alters the expression and phosphorylation patterns of mice hepatic proteome, which possibly attributes to considerably different sets of rhythmic candidates compared to wild-type. It is, therefore, reasonable to accentuate that circadian rhythms are not obliterated in mammalian systems due to deletion of the core clock genes. Overall design: Wild-type and Bmal1-/- MSF cells were grown in Dulbecco's Modified Eagle Medium (DMEM) containing 10% (v/v) HyClone III Serum (Analab; Cat # SH30109.03), 1/100  Glutamax-I (Invitrogen; Cat # 35050-038), 1/100 Penicillin-Streptomycin (SIGMA; Cat # P4333) and 1/500 MycoZap™ (Lonza; Cat # VZA-2022) in multiple six-well plates until complete confluency (n=3, per time-point, per genotype). The confluent MSF cells were treated with 100nM (final concentration) of dexamethasone (DEX) for 15 min to synchronize the cells [Balsalobre et al. Science 2000]29. MSF cells were then washed three times with PBS (37°C) and were incubated in HEPES-buffered Medium; 1X DMEM powder (SIGMA; Cat # D5030), 0.35mg/mL sodium bicarbonate, 5mg/mL glucose, 0.01M HEPES, 10% (v/v) HyClone III Serum, 1/100 Glutamax-I, and 1/500 MycoZap™, pH 7.3 (adjusted with HCl) and osmolality 320 mOsm (adjusted with NaCl) at 37°C under DD cycle. 48 hrs after DEX treatment, MSF cells were harvested at every three or two hours interval for three days for transcriptomics analyses.