Novel Innate Immune Genes Regulating the Macrophage Response to Gram Positive Bacteria
Novel Innate Immune Genes Regulating the Macrophage Response to Gram Positive Bacteria
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DOI：
10.1534/genetics.115.185314
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发表时间：
2016-09-01
期刊：
影响因子：
3.3
通讯作者：
Yang, Ivana V.
中科院分区：
文献类型：
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作者：
Alper, Scott;Warg, Laura A.;Yang, Ivana V.
宿主toll样受体和其他先天免疫信号分子的变异改变了感染易感性。然而，在先天免疫反应中观察到的变异性只有一部分是由这些途径中的已知基因所解释的。因此，鉴定其他基因调节对革兰氏阳性细菌的反应是必要的。用脂科技酸（LTA）刺激43种自交系小鼠骨髓源性巨噬细胞（BMMs），脂科技酸是革兰氏阳性细菌细胞壁的主要成分。测定促炎细胞因子IL-6、IL-12和TNF-的浓度。利用细胞因子反应进行全基因组关联（WGA）定位，然后进行网络分析，确定候选基因的优先级。为了确定哪些候选基因可能负责调节LTA反应，我们使用RNA干扰（RNAi）抑制候选基因，并在RAW264.7巨噬细胞中过表达。Bdkrb1基因缺失小鼠的bmm被用来评估Bdkrb1基因缺失对LTA、热死肺炎链球菌和热死金黄色葡萄球菌反应的影响。WGA图谱鉴定出117个基因座：IL-6分析鉴定出20个基因座（平均基因座大小0.133 Mb， 18个基因），IL-12分析鉴定出5个基因座（平均0.201 Mb， 7个基因），TNF-分析鉴定出92个基因座（平均0.464 Mb， 186个基因，其中46个通过网络分析优先排序）。随后对71个靶基因进行小干扰RNA筛选，发现4个基因（Bdkrb1、Blnk、Fbxo17和Nkx6-1）在LTA刺激后受到抑制，导致细胞因子的产生显著减少。这四个基因的过度表达导致LTA应答中细胞因子的产生显著增加。缺乏bdkrb1的巨噬细胞对LTA和热杀伤金黄色葡萄球菌的反应较低，验证了遗传和RNAi方法鉴定对LTA反应的新调节因子。我们已经确定了四个先天免疫反应基因，可能有助于革兰氏阳性细菌的敏感性。
Host variation in Toll-like receptors and other innate immune signaling molecules alters infection susceptibility. However, only a portion of the variability observed in the innate immune response is accounted for by known genes in these pathways. Thus, the identification of additional genes that regulate the response to Gram positive bacteria is warranted. Bone marrow-derived macrophages (BMMs) from 43 inbred mouse strains were stimulated with lipotechoic acid (LTA), a major component of the Gram positive bacterial cell wall. Concentrations of the proinflammatory cytokines IL-6, IL-12, and TNF- were measured. In silico whole genome association (WGA) mapping was performed using cytokine responses followed by network analysis to prioritize candidate genes. To determine which candidate genes could be responsible for regulating the LTA response, candidate genes were inhibited using RNA interference (RNAi) and were overexpressed in RAW264.7 macrophages. BMMs from Bdkrb1-deficient mice were used to assess the effect of Bdkrb1 gene deletion on the response to LTA, heat-killed Streptococcus pneumoniae, and heat-killed Staphylococcus aureus. WGA mapping identified 117 loci: IL-6 analysis yielded 20 loci (average locus size = 0.133 Mb; 18 genes), IL-12 analysis produced 5 loci (0.201 Mb average; 7 genes), and TNF- analysis yielded 92 loci (0.464 Mb average; 186 genes of which 46 were prioritized by network analysis). The follow-up small interfering RNA screen of 71 target genes identified four genes (Bdkrb1, Blnk, Fbxo17, and Nkx6-1) whose inhibition resulted in significantly reduced cytokine production following LTA stimulation. Overexpression of these four genes resulted in significantly increased cytokine production in response to LTA. Bdkrb1-deficient macrophages were less responsive to LTA and heat-killed S. aureus, validating the genetic and RNAi approach to identify novel regulators of the response to LTA. We have identified four innate immune response genes that may contribute to Gram positive bacterial susceptibility.