Viruses exploit phosphorylation of both viral and host proteins to support viral replication. In this study, we demonstrate that porcine reproductive and respiratory syndrome virus replicase nsp2, and two nsp2-related −2/-1 frameshifting products, nsp2TF and nsp2N, are hyper-phosphorylated. By mapping phosphorylation sites, we subdivide an extended, previously uncharacterized region, located between the papain-like protease-2 (PLP2) domain and frameshifting site, into three distinct domains. These domains include two large hypervariable regions (HVR) with putative intrinsically disordered structures, separated by a conserved and partly structured interval domain that we defined as the inter-HVR conserved domain (IHCD). Abolishing phosphorylation of the inter-species conserved residue serine⁹¹⁸, which is located within the IHCD region, abrogates accumulation of viral genomic and subgenomic RNAs and recombinant virus production. Our study reveals the biological significance of phosphorylation events in nsp2-related proteins, emphasizes pleiotropic functions of nsp2-related proteins in the viral life cycle, and presents potential links to pathogenesis.

病毒利用病毒和宿主蛋白的磷酸化来支持病毒复制。在这项研究中，我们证明了猪生殖和呼吸综合症病毒复制酶nsp2和两个与nsp2相关的-2 / -1移码产物nsp2TF和nsp2N被过度磷酸化。通过定位磷酸化位点，我们将位于木瓜蛋白酶样蛋白酶2（PLP2）域和移码位点之间的扩展的，以前未表征的区域细分为三个不同的域。这些域包括两个具有假定的固有无序结构的大高变区（HVR），这些区域由一个保守且部分结构化的间隔域隔开，我们将其定义为HVR间保守域（IHCD）。消除种间保守残基丝氨酸^918的磷酸化位于IHCD区域内的cDNA消除了病毒基因组和亚基因组RNA的积累以及重组病毒的产生。我们的研究揭示了nsp2相关蛋白中磷酸化事件的生物学意义，强调了nsp2相关蛋白在病毒生命周期中的多效性功能，并提出了与发病机理的潜在联系。