The lipopeptide Pam3CSK4 inhibits Rift Valley fever virus infection and protects from encephalitis

Rift Valley fever virus (RVFV) is an encephalitic bunyavirus that can infect neurons in the brain. There are no approved therapeutics that can protect from RVFV encephalitis. Innate immunity, the first line of defense against infection, canonically antagonizes viruses through interferon signaling. We found that interferons did not efficiently protect primary cortical neurons from RVFV, unlike other cell types. To identify alternative neuronal antiviral pathways, we screened innate immune ligands and discovered that the TLR2 ligand Pam3CSK4 inhibited RVFV infection, and other bunyaviruses. Mechanistically, we found that Pam3CSK4 blocks viral fusion, independent of TLR2. In a mouse model of RVFV encephalitis, Pam3CSK4 treatment protected animals from infection and mortality. Overall, Pam3CSK4 is a bunyavirus fusion inhibitor active in primary neurons and the brain, representing a new approach toward the development of treatments for encephalitic bunyavirus infections. To define the neuronal response to TLR2 stimulation, we performed RNAseq on rat cortical neurons (7 days ex vivo) stimulated with vehicle, Pam3CSK4 (10ug/mL), or Pam2CSK4 (10ug/mL) for 6 hours. Three replicates per condition. We then used differential gene analysis to compare genes induced by Pam3CSK4 or Pam2CSK4 relative to vehicle

裂谷热病毒（Rift Valley fever virus, RVFV）是一类可侵染大脑神经元的嗜脑性布尼亚病毒。目前尚无获批的可用于预防RVFV脑炎的治疗药物。固有免疫作为机体抵御感染的第一道防线，经典地通过干扰素信号通路发挥病毒拮抗作用。本研究发现，与其他细胞类型不同，干扰素无法有效保护原代皮层神经元免受RVFV感染。为鉴定神经元内的替代性抗病毒通路，我们对固有免疫配体进行了筛选，发现Toll样受体2（Toll-like receptor 2, TLR2）配体Pam3CSK4可抑制RVFV及其他布尼亚病毒的感染。机制研究表明，Pam3CSK4可独立于TLR2阻断病毒融合过程。在RVFV脑炎小鼠模型中，Pam3CSK4给药可使动物免受感染并降低死亡率。综上，Pam3CSK4是一种可在原代神经元及脑组织中发挥活性的布尼亚病毒融合抑制剂，为嗜脑性布尼亚病毒感染的治疗开发提供了全新策略。为明确神经元对TLR2刺激的转录应答，我们对经溶剂对照、10μg/mL Pam3CSK4或10μg/mL Pam2CSK4处理6小时的体外培养7天的大鼠皮层神经元进行了RNA测序（RNA sequencing, RNA-seq），每组设置3个生物学重复。随后我们通过差异基因表达分析，对比了Pam3CSK4或Pam2CSK4相较于溶剂对照所诱导的基因表达差异。