The cGAS-STING pathway, orchestrating complicated transcriptome-wide immune responses, is essential for host antiviral defense but can also drive immunopathology in severe COVID-19. Here, we performed time-course RNA-Seq experiments to dissect the transcriptome expression dynamics at the gene-isoform level after cGAS-STING pathway activation. The in-depth time-course transcriptome after cGAS-STING pathway activation within 12 hours enabled quantification of 48,685 gene isoforms. By employing regression models, we obtained 13,232 gene isoforms with expression patterns significantly associated with the process of cGAS-STING pathway activation, which were named activation-associated isoforms. The combination of hierarchical and k-means clustering algorithms revealed four major expression patterns of activation-associated isoforms, including two clusters with increased expression patterns enriched in cell cycle, autophagy, antiviral innate-immune functions, and COVID-19 coronavirus disease pathway, and two clusters showing decreased expression pattern that mainly involved in ncRNA metabolism, translation process, and mRNA processing. Importantly, by merging four clusters of activation-associated isoforms, we identified three types of genes that underwent isoform usage alteration during the cGAS-STING pathway activation. We further found that genes exhibiting protein-coding and non-protein-coding gene isoform usage alteration were strongly enriched for the factors involved in innate immunity and RNA splicing. Notably, overexpression of an enriched splicing factor, EFTUD2, shifted transcriptome towards the cGAS-STING pathway activated status and promoted protein-coding isoform abundance of several key regulators of the cGAS-STING pathway. Taken together, our results revealed the isoform-level gene expression dynamics of the cGAS-STING pathway and uncovered novel roles of splicing factors in regulating cGAS-STING pathway mediated immune responses.

cGAS-STING 通路协调复杂的全转录组免疫反应，对于宿主抗病毒防御至关重要，但也可以驱动严重 COVID-19 的免疫病理学。在这里，我们进行了时程 RNA-Seq 实验，以剖析 cGAS-STING 通路激活后基因同种型水平的转录组表达动力学。cGAS-STING 通路激活后 12 小时内的深度时程转录组能够对 48,685 种基因亚型进行量化。通过使用回归模型，我们获得了 13,232 个基因亚型，其表达模式与 cGAS-STING 通路激活过程显着相关，这些亚型被命名为激活相关亚型。分层和 k 均值聚类算法的组合揭示了激活相关亚型的四种主要表达模式，包括在细胞周期、自噬、抗病毒先天免疫功能和 COVID-19 冠状病毒疾病途径中表达模式增加的两个簇，以及主要涉及 ncRNA 代谢、翻译过程和 mRNA 加工的表达模式减少的两个簇。重要的是，通过合并四个激活相关异构体簇，我们确定了三种类型的基因，这些基因在 cGAS-STING 通路激活期间经历了异构体使用改变。我们进一步发现，表现出蛋白质编码和非蛋白质编码基因亚型使用改变的基因在先天免疫和 RNA 剪接中所涉及的因素中得到了极大的丰富。值得注意的是，富集剪接因子 EFTUD2 的过度表达，将转录组转向 cGAS-STING 通路激活状态，并促进 cGAS-STING 通路几个关键调节因子的蛋白质编码亚型丰度。综上所述，我们的结果揭示了 cGAS-STING 通路的亚型水平基因表达动力学，并揭示了剪接因子在调节 cGAS-STING 通路介导的免疫反应中的新作用。