Diurnal liver transcriptome in wt and nono-/- mice

The liver circadian transcriptome results from the combined action of a circadian clock and feeding/fasting rhythms. Since we found that (1) the number of NONO-containing speckles increases in response to feeding, (2) NONO interacts with RNA processing factors and (3) it binds to the introns of a large number of protein coding genes, we hypothesized that NONO may contribute to the daily rhythm of hepatic gene expression. To address whether NONO contributes to rhythmic gene expression in response to feeding/fasting, diurnal liver transcriptomes were assessed in nono-/- and wt littermate mice fed a normal diet during the night time (ZT12-24). We habituated mice to a 12:12 LD cycle with food available only at lights off (ZT12-24) for one week and in the second week collected mice every 2h throughout the 24h day. Both total and nuclear RNAs were sequenced after ribosomal RNA depletion Livers were collected at 12 different time points during a 24h day (ZT0-ZT22). For each liver sample, total and nuclear RNA were extracted for a total of 48 samples. Each sample is a pool of 2 mice.肝脏昼夜转录组的产生源于昼夜节律钟与摄食/禁食节律的共同调控作用。鉴于本研究发现：① 摄食信号可诱导含NONO的核斑数量增加；② NONO能够与RNA加工因子发生相互作用；③ NONO可结合大量蛋白编码基因的内含子区域，因此我们提出假说：NONO可能参与调控肝脏基因表达的昼夜节律。为探究NONO是否参与响应摄食/禁食节律的基因表达调控，我们对夜间（ZT12-24，即关灯时段）喂食正常饲料的nono基因敲除（nono-/-）与野生型（wild type，wt）同窝小鼠的肝脏昼夜转录组进行了检测。实验前，我们先让小鼠适应12:12光暗（Light-Dark，LD）循环环境，且仅在关灯时段（ZT12-24）提供食物，持续一周；实验第二周内，我们每2小时采集一次小鼠样本，覆盖完整24小时周期。我们对核糖体RNA进行去除后，分别对总RNA与核RNA进行了测序。本实验在24小时周期内的12个不同时间点（ZT0至ZT22）采集肝脏样本。每个肝脏样本均提取总RNA与核RNA，总计获取48份样本；每份样本由2只小鼠的肝脏混合制备而成。