Differential Expression of Immune Related Genes in Taste Buds of Fed and Fasted Mice. Differential Expression of Immune Related Genes in Taste Buds of Fed and Fasted Mice

Purpose: The goal of this study is to compare the taste bud transcriptome of fasted and fed mice. By doing this we hope to gain more information about the taste bud transcriptome as well as determine transcriptional changes that may be linked with pre or post-ingestive states. Methods: Whole taste bud mRNA profiles of WT adult C57BL/6J mice were generated by Illumina HiSeq 2000 single end RNA sequencing. Prior to taste bud RNA isolation, mice were either food-deprived, fed, or given ad libitum access to standard rodent chow as a control. Sequence read quality was assesed using FastQC, and adapter sequences were trimmed using trimmomatic. Reads were then aligned to the GRCm38.p4 reference genome using Tophat2 followed by Cufflinks analysis, then gene level differential expression was conducted using Cuffdiff, and visualized via cummeRbund. Gene enrichment analysis of differentially expressed genes was performed using DAVID and Panther Gene Ontology. Results: We mapped about 30 million sequence reads per sample to the mouse genome, build GRCm38.p4, and identified 144 differentially expressed genes (FDR of 5%) between the taste buds of food-restricted, fed, and ad libitum control mice. Gene enrichment analysis of differentially expressed genes showed enrichment of pathways associated with cytokines, immunity, cytoskeletal structure, chaperone proteins, and protease inhibitors. Conclusions: Our study represents a detailed analysis of the murine taste bud transcriptome generated using RNA-seq technology. Our results highlight cellular pathways that may be differentially regulated in taste receptor cells during different pre/post-ingestive states. Additionally, as limited transcriptome information is available for taste buds, this dataset can serve as a resource for the discovery of genes novel to the taste bud. Overall design: Whole taste bud mRNA profiles of 20-24 week old WT C57B/6J (fed, fasted, or ad libitum fed) were generated by RNA sequencing using Illumina HiSeq 2000.

研究目的：本研究旨在对比禁食与进食小鼠的味蕾转录组，以期获取更多味蕾转录组相关信息，并明确可能与摄食前/摄食后状态相关的转录水平变化。
实验方法：采用因美纳（Illumina）HiSeq 2000单端RNA测序（RNA-seq）技术，获取野生型（Wild Type, WT）成年C57BL/6J小鼠的全味蕾mRNA表达谱。在分离味蕾RNA前，将小鼠分为三组：食物剥夺组、进食组，以及自由采食标准啮齿类饲料的对照组。使用FastQC评估测序读段质量，通过Trimmomatic去除接头序列。随后利用Tophat2将读段比对至GRCm38.p4参考基因组，再经Cufflinks分析；采用Cuffdiff进行基因水平差异表达分析，并通过cummeRbund完成可视化。使用DAVID与Panther基因本体（Gene Ontology, GO）数据库对差异表达基因进行基因富集分析。
实验结果：每个样本约有3000万条测序读段被比对至小鼠GRCm38.p4参考基因组，在食物限制组、进食组与自由采食对照组的味蕾中，共鉴定出144个差异表达基因（错误发现率（False Discovery Rate, FDR）为5%）。对差异表达基因的富集分析显示，其显著富集于细胞因子、免疫、细胞骨架结构、分子伴侣蛋白以及蛋白酶抑制剂相关的通路。
研究结论：本研究对基于RNA测序技术获得的小鼠味蕾转录组进行了详尽分析。研究结果揭示了在不同摄食前/摄食后状态下，味觉受体细胞中可能存在差异调控的细胞通路。此外，当前味蕾转录组的相关信息较为有限，本数据集可作为味蕾新基因发现的研究资源。
实验整体设计：采用因美纳（Illumina）HiSeq 2000 RNA测序技术，获取20~24周龄野生型C57BL/6J小鼠（分为进食组、禁食组与自由采食组）的全味蕾mRNA表达谱。