Differential gene regulation downstream of murine Tmem173 variants

We report positional cloning and characterization of a novel Sting allele that fails to activate IFN production in the wild-derived mice of MOLF/Ei strain in response to HSV (Herpes Simplex Virus) and Listeria monocytogenes both in vitro and in vivo. We show that previously uncharacterized mutations in the N-terminal of STING are responsible for low levels of IFN due to failure of MOLF STING to respond to cytosolic STING agonists such as 2'3'cGAMP, 5,6-Dimethylxanthenone-4-acetic acid (DMXAA) or poly(deoxyadenylic-deoxythymidylic) acid (dAdT). Using Next-Generation Sequencing (NGS) for the analysis of DNA-responses in congenic C57BL6.StingMOLF/MOLF (MOLF STING) mouse macrophages, we show that these mutations in MOLF/Ei discriminate in responses between different STING agonists. Macrophages from C57BL6.StingB6/B6 (B6 STING), or B6 STING expressing littermates were stimulated as a positive control for STING activation, and STING -/- (STING KO) macrophages served as a negative control. To examine differential gene regulation downstream of murine Tmem173 (STING) allelic variants. We stimulated macrophages from B6 mice congenic for MOLF STING; macrophages from B6 STING expressing littermates were stimulated as a positive control for STING activation, and STING -/- (STING KO) macrophages served as a negative control. One replicate of RNA-seq reads after each stimulation provided. Conditions are peritoneal macrophages from B6 congenic mice expressing: B6 STING, MOLF STING, or STING KO stimulated with 2'3'cGAMP, 5,6-Dimethylxanthenone-4-acetic acid (DMXAA), poly(deoxyadenylic-deoxythymidylic) acid (dAdT), or no treatment. This constitutes a total of 12 reads analyzed in this data set.本研究报道了一种新型干扰素基因刺激因子（STING，Sting）等位基因的位置克隆与功能表征：该等位基因在MOLF/Ei品系野生来源小鼠的体内外环境中，均无法在单纯疱疹病毒（Herpes Simplex Virus，HSV）与单核细胞增生李斯特菌（Listeria monocytogenes）的刺激下激活干扰素（IFN）的产生。研究发现，STING N端此前未被表征的突变，是导致MOLF品系STING无法响应胞质STING激动剂（如2'3'-cGAMP、5,6-二甲基占吨酮-4-乙酸（DMXAA）以及聚脱氧腺苷-聚脱氧胸腺苷酸（dAdT）），进而引发干扰素产生水平低下的核心原因。本研究利用下一代测序（Next-Generation Sequencing，NGS）技术，对同类系C57BL/6.StingMOLF/MOLF（MOLF STING）小鼠巨噬细胞的DNA应答反应进行分析，结果表明MOLF/Ei品系的上述突变可区分不同STING激动剂所诱导的细胞应答。实验中，以表达C57BL/6.StingB6/B6（B6 STING）或B6 STING的同窝小鼠巨噬细胞作为STING激活的阳性对照，以STING基因敲除（STING -/-，即STING KO）巨噬细胞作为阴性对照。为探究小鼠Tmem173（STING）等位基因变异下游的差异基因调控机制，我们对携带MOLF STING的同类系B6小鼠的巨噬细胞进行刺激；同时以表达B6 STING的同窝小鼠巨噬细胞作为STING激活的阳性对照，以STING基因敲除巨噬细胞作为阴性对照。本数据集提供了每种刺激条件下的一组RNA测序（RNA-seq）读段重复数据。实验分组包括：分别经2'3'-cGAMP、5,6-二甲基占吨酮-4-乙酸（DMXAA）、聚脱氧腺苷-聚脱氧胸腺苷酸（dAdT）刺激，或未接受任何处理的、表达B6 STING、MOLF STING或STING基因敲除的同类系B6小鼠腹腔巨噬细胞。本数据集共分析了12组读段数据。