RNA-Sequence Analysis of Primary Alveolar Macrophages after In Vitro Infection with Porcine Reproductive and Respiratory Syndrome Virus Strains of Differing Virulence

Porcine reproductive and respiratory syndrome virus (PRRSV) mainly infects porcine alveolar macrophages (PAMs), resulting in porcine reproductive and respiratory syndrome (PRRS) in pigs. Most of the transcriptomic studies on PAMs infected with PRRSV conducted thus far have made use of microarray technology. Here, we investigated the transcriptome of PAMs in vitro at 12 h post-infection with two European PRRSV strains characterized by low (Lelystad, LV) and high (Lena) virulence through RNA-Seq. The expression levels of genes, isoforms, alternative transcription start sites (TSS) and differential promoter usage revealed a complex pattern of transcriptional and post-transcriptional gene regulation upon infection with the two strains. Gene ontology analysis confirmed that infection of PAMs with both the Lena and LV strains affected signaling pathways directly linked to the innate immune response, including interferon regulatory factors (IRF), RIG1-like receptors, TLRs and PKR pathways. The results confirmed that interferon signaling is crucial for transcriptional regulation during PAM infection. IFN-β1 and IFN-αω, but not IFN-α, were up-regulated following infection with either the LV or Lena strain. The down-regulation of canonical pathways, such as the interplay between the innate and adaptive immune responses, cell death and TLR3/TLR7 signaling, was observed for both strains, but Lena triggered a stronger down-regulation than LV. This analysis contributes to a better understanding of the interactions between PRRSV and PAMs and outlines the differences in the responses of PAMs to strains with different levels of virulence, which may lead to the development of new PRRSV control strategies.

猪繁殖与呼吸综合征病毒（Porcine reproductive and respiratory syndrome virus, PRRSV）主要感染猪肺泡巨噬细胞（porcine alveolar macrophages, PAMs），可引发猪群罹患猪繁殖与呼吸综合征（Porcine reproductive and respiratory syndrome, PRRS）。迄今为止，绝大多数针对PRRSV感染PAMs的转录组学研究均采用微阵列技术开展。本研究借助RNA测序（RNA-Seq）技术，分析了体外培养的PAMs在感染两种欧洲PRRSV毒株（分别为低毒力株Lelystad（LV）与高毒力株Lena）后12小时的转录组特征。研究结果显示，感染两类毒株后，基因、剪接异构体、可变转录起始位点（alternative transcription start sites, TSS）的表达水平以及启动子使用差异，均呈现出复杂的转录及转录后基因调控模式。基因本体分析（Gene Ontology, GO）证实，Lena与LV两株病毒感染PAMs后，均会影响与先天免疫应答直接相关的信号通路，包括干扰素调节因子（interferon regulatory factors, IRF）通路、RIG1样受体（RIG1-like receptors）通路、Toll样受体（TLRs）通路以及蛋白激酶R（PKR）通路。本研究结果证实，干扰素信号通路在PAMs感染过程的转录调控中发挥关键作用。感染LV或Lena毒株后，IFN-β1与IFN-αω的表达均会上调，但IFN-α的表达无显著变化。两类毒株均会导致多条经典通路出现下调，包括先天与适应性免疫应答互作、细胞死亡以及TLR3/TLR7信号通路，且Lena毒株引发的下调程度显著强于LV毒株。本研究有助于进一步解析PRRSV与PAMs之间的相互作用，并阐明PAMs对不同毒力毒株的应答差异，可为新型PRRSV防控策略的开发提供理论支撑。