Human 5-lipoxygenase regulates transcription by association to euchromatin [FAIRE-seq]

Human 5-lipoxygenase (5-LO) is the key enzyme of leukotriene biosynthesis, mostly expressed in leukocytes and thus a crucial component of the innate immune system. In this study, we show that 5-LO, beside its canonical function as a metabolizing enzyme, is also a regulator of gene expression in the monocytic cell line Mono Mac 6 (MM6). Using RNA-Seq, we identified 5-LO regulated genes which could be clustered to several biological functions, e.g. immune/defense response, cell adhesion, transcription and growth/developmental processes. Thereby, the majority of genes were upregulated by 5-LO KO. By conducting a ChIP-Seq analysis, we proved an association of 5-LO to DNA. In summary, we demonstrate an additional noncanonical function of 5-LO as a direct transcriptional regulator in MM6 cells that presumably regulates specific genes in dependence on histone acetylation. Overall design: Three independent replicates of Formaldehyde Assisted Isolation of Regulatory Elemens (FAIRE)-Seq from MonoMac 6 cells (MM6) with and without 5-Lipoxygenase knockout after 72h treatment with TGF-ß/1,25(OH)2D3 and with or without 2h LPS stimulation.

人类5-脂氧合酶（5-lipoxygenase, 5-LO）是白三烯生物合成的关键酶，主要表达于白细胞中，因此是先天免疫系统的重要组成部分。
在本研究中，我们证实5-LO除了作为代谢酶发挥经典功能外，还可调控单核细胞系Mono Mac 6（MM6）中的基因表达。通过RNA测序（RNA-Seq）技术，我们鉴定出受5-LO调控的基因，这些基因可聚类为多种生物学功能，例如免疫/防御反应、细胞黏附、转录及生长发育过程。其中大多数基因经5-LO敲除后表达上调。通过染色质免疫沉淀测序（ChIP-Seq）分析，我们证实5-LO可与DNA结合。
综上，我们证明了5-LO在MM6细胞中存在一种额外的非经典功能，即作为直接转录调控因子，其可能依赖于组蛋白乙酰化调控特定基因的表达。
整体实验设计：针对经转化生长因子-β/1,25-二羟基维生素D3处理72小时、并施以或不施以2小时脂多糖（Lipopolysaccharide, LPS）刺激的MonoMac 6（MM6）细胞（含5-脂氧合酶敲除组与野生型对照组），每个处理条件均设置3次独立生物学重复，随后开展甲醛辅助调控元件分离测序（FAIRE-Seq）。