Muscle GR and BMAL1 epigenomic binding at ZT4 after vehicle versus prednisone pulse at ZT0

Exogenous glucocorticoids interact with the circadian clock, but little attention is paid to their time of intake. Here we investigated the epigenetic effects of prednisone in muscle when pulsed as single 1mg/kg i.p. dose in mice WT mice at ZT0, i.e. when endogenous corticosterone is low. At 4-hours post-injection (ZT4), we performed ChIP-seq analysis in muscle tissue for the glucocorticoid receptor (GR), as well as BMAL1, clock factor active in the light-phase. We also performed ChIP-seq for RNApol-II to quantitate changes in RNApol-II recruitment. We repeated the GR and RNApol-II analyses in muscles from BMAL1-knockout mice. 3 samples BMAL-WT muscle + vehicle; 3 samples BMAL-WT muscle + prednisone; 3 samples BMAL-KO muscle + vehicle; 3 samples BMAL-KO muscle + prednisone. Each sample was ~10 ng immunoprecipitated chromatin from 2 pooled quadriceps muscles of one mouse. Primary antibodies: rabbit polyclonal anti-GR (ABclonal #A2164), rabbit polyclonal anti-RNApol-II (A11181) and mouse monoclonal anti-BMAL1 (DSHB # PCRP-ARNTL-1C12).