Determination of Lsd1 function in brown adipose tissue by global transcriptome analysis [RNA-seq]

To investigate the role of Lsd1 in BAT, we crossed mice harboring conditional Lsd1 alleles (Zhu et al., 2014) with the Ucp1-Cre deleter strain (Turpin et al., 2014), which results in Cre-mediated loss of Lsd1 selectively in brown adipocytes. Transciptome analysis of BAT obtained from control and knock-out mice revealed 3322 genes differentially expressed between Lsd1 knock-out and control mice at 10 weeks of age (p-value<10-2). We selectively knocked-out Lsd1 in brown adipose tissue by crossing Lsd1 floxed mice to Ucp1-Cre mice. RNA from BAT of 10 week.old control (Lsd1 floxed mice) and knock-out mice was extracted with trizol according to the manufacturer protocol. RNA samples were sequenced by the standard Illumina protocol to create raw sequence files (.fastq files). We annotated these reads to the mm10 build of the mouse genome using TopHat version 2. The aligned reads were counted with the homer software (analyze RNA) and DEG's were identified using EdgeR and DESeq version 1.8.3. LSD1 binding profile in fat cells - samples and experiments will continue to be added to this study over the course of the project.为探究赖氨酸特异性去甲基化酶1（Lsd1）在棕色脂肪组织（BAT, brown adipose tissue）中的功能，我们将携带条件性Lsd1等位基因的小鼠（Zhu等，2014）与Ucp1-Cre删除工具株（Turpin等，2014）进行杂交，实现了Cre重组酶介导的棕色脂肪细胞内Lsd1的特异性敲除。对对照小鼠与敲除小鼠的棕色脂肪组织开展转录组分析后发现，小鼠10周龄时，Lsd1敲除组与对照组间共有3322个基因呈现差异表达（p值<10^-2）。为开展本研究，我们通过将Lsd1 floxed小鼠与Ucp1-Cre小鼠杂交，特异性敲除小鼠棕色脂肪组织中的Lsd1。按照厂商提供的实验规程，使用Trizol试剂提取10周龄对照小鼠（仅携带Lsd1 floxed等位基因）与敲除小鼠的棕色脂肪组织总RNA。随后依照标准Illumina测序流程对RNA样本进行测序，获得原始测序文件（.fastq文件）。我们使用TopHat 2版本将测序读段比对至小鼠基因组mm10版本，借助homer软件（analyze RNA工具）对比对后的读段进行计数，并通过EdgeR与DESeq 1.8.3版本鉴定差异表达基因（DEGs, differentially expressed genes）。本研究中脂肪细胞的LSD1结合谱相关样本与实验数据，将在项目推进过程中持续补充至本数据集。