Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, OK, 73190, USA.
Sci Rep. 2018 Nov 16;8(1):16931. doi: 10.1038/s41598-018-35184-y.
CREB and C/EBP β signaling pathways are modulated during inflammation and also targeted by Bacillus anthracis edema toxin (ET), but how these factors individually and jointly contribute to changes in immune cell function is poorly understood. Using CRISPR/Cas9 gene editing, macrophage cell lines lacking CREB and isoforms of C/EBP β were generated and analyzed for changes in responses to LPS, ET, and IL-4. Macrophages lacking C/EBP β suppressed induction of IL-10 and Arg1, while IL-6 was increased in these cells following exposure to LPS. Examination of C/EBP β isoforms indicated the 38 kDa isoform was necessary for the expression of IL-10 and Arg1. ChIP-Seq analysis of CREB and C/EBP β binding to targets on the chromosome of human PBMC identified several regions where both factors overlapped in their binding, suggesting similar gene targeting or cooperative effects. Based on the ChIP-Seq data, a panel of previously unknown targets of CREB and C/EBP β was identified and includes genes such as VNN2, GINS4, CTNNBL1, and SULF2. Isoforms of a transcriptional corepressor, transducin-like enhancer of Split (TLE), were also found to have CREB and C/EBP β binding their promoter and were up regulated by ET. Finally, we explore a possible layer of C/EBP β regulation by a protein complex consisting of adenomatous polyposis coli (APC) and PKA. Collectively, these data provide new insights into the role of CREB and C/EBP β as immunosignaling regulators and targets of an important bacterial virulence factor.
CREB 和 C/EBPβ 信号通路在炎症过程中受到调节，炭疽杆菌水肿毒素 (ET) 也靶向这些通路，但这些因素如何单独和共同导致免疫细胞功能的变化还知之甚少。使用 CRISPR/Cas9 基因编辑，生成了缺乏 CREB 和 C/EBPβ 同工型的巨噬细胞系，并分析了它们对 LPS、ET 和 IL-4 的反应变化。缺乏 C/EBPβ 的巨噬细胞抑制了 IL-10 和 Arg1 的诱导，而这些细胞在暴露于 LPS 后 IL-6 增加。对 C/EBPβ 同工型的检查表明，38 kDa 同工型是 IL-10 和 Arg1 表达所必需的。对 CREB 和 C/EBPβ 结合到人类 PBMC 染色体上靶标的 ChIP-Seq 分析确定了几个因素重叠结合的区域，表明存在类似的基因靶向或协同作用。基于 ChIP-Seq 数据，鉴定了一组以前未知的 CREB 和 C/EBPβ 靶点，包括 VNN2、GINS4、CTNNBL1 和 SULF2 等基因。转录核心抑制因子 transducin-like enhancer of Split (TLE) 的同工型也被发现具有 CREB 和 C/EBPβ 结合其启动子的能力，并被 ET 上调。最后，我们探索了由腺瘤性结肠息肉病 (APC) 和 PKA 组成的蛋白质复合物对 C/EBPβ 调节的可能层。总之，这些数据为 CREB 和 C/EBPβ 作为免疫信号调节因子以及一个重要细菌毒力因子的靶标提供了新的见解。
