Transcriptomic analysis of cortex tissue from neuron-specific Bmal1 KO mice

The circadian clock protein Bmal1 is critical for maintain circadian transcriptional function and rhythms. Global deletion of Bmal1 renders whole mice behaviorally arrhythmic. However, cell-specific deletion of Bmal1 can reveal specific transcripts regulated by Bmal1 in a cell-specific manner. Here we used a BAC-transgenic Camk2a-iCre line to delete Bmal1 in a pan-neuronal manner. This mouse has previously been shown to have widespread Bmal1 deletion in neurons and to be arrhythmic (PMID: 25525750). We performed bulk RNAseq on cerebral cortex tissue from Camk2a-iCre+;Bmal1(fx/fx) and Cre- control littermates. Appropriate Bmal1 targets were disregulated as expected in these mice. Pathways analysis revealed transcripts involved in Parkinson Disease and oxidative phosphorylation to be enriched. Camk2a-iCre+;Bmal1(fx/fx) and Cre-;Bmal1(fx/fx) control mice N=3/genotype, were kept under normal 12:12 light:dark conditions until 4 months of age, then anesthetized and perfused with PBS. Cerebral cortex tissue was dissected and frozen, then later subjected to bulk RNAseq analysis. These Camk2a-iCre mice are BAG transgenic as previously described (see Izuma et al PMID: 25525750), which have pan-neuronal Bmal1 deletion and circadian disruption.生物钟核心蛋白Bmal1对于维持生物钟转录调控功能与生物节律至关重要。全身性敲除Bmal1会使小鼠整体表现出行为节律紊乱。然而，细胞特异性敲除Bmal1则可揭示不同细胞类型中受Bmal1调控的特异性转录本。本研究采用细菌人工染色体（BAC）转基因的Camk2a-iCre工具鼠，以泛神经元特异性方式敲除Bmal1。此前已有研究证实，该工具鼠的神经元中广泛存在Bmal1敲除，且表现出节律紊乱（PMID: 25525750）。我们对Camk2a-iCre+;Bmal1(fx/fx)小鼠及其Cre-同窝对照小鼠的大脑皮层组织开展了批量RNA测序（bulk RNAseq）。该组小鼠中，预期的Bmal1靶基因均出现表达失调。通路富集分析显示，与帕金森病（Parkinson Disease）和氧化磷酸化相关的转录本呈现富集趋势。本研究中，Camk2a-iCre+;Bmal1(fx/fx)小鼠与Cre-;Bmal1(fx/fx)对照小鼠每组各3只（N=3/基因型），均在标准12小时光照:12小时黑暗的光照条件下饲养至4月龄，随后经麻醉后用磷酸盐缓冲液（PBS）灌注取材。分离大脑皮层组织并速冻保存，后续进行批量RNA测序分析。本研究使用的Camk2a-iCre小鼠为细菌人工染色体转基因小鼠，如此前文献所述（参见Izuma等人，PMID: 25525750），该类小鼠存在泛神经元Bmal1敲除与生物钟节律紊乱。