Lactate-lactylation-HSPA6 axis promotes PRRSV replication by impairing IFN-β production

Lactate, traditionally considered a metabolic by-product, has recently been identified as a substrate for the induction of lactylation, a newly identified epigenetic modification that plays an important role in the regulation of host gene expression. Our previous study showed that lactate levels were significantly elevated in cells infected with the porcine reproductive and respiratory syndrome virus (PRRSV), an Arterivirus that has devastated the swine industry worldwide for over 30 years. However, the role of elevated lactate in PRRSV infections remains unknown. In this study, we found that lactate was required for optimal PRRSV proliferation, and PRRSV infection increased cellular lactylation in a dose-dependent manner. Using the Cleavage Under Targets and Tagmentation (CUT&Tag) combined with RNA sequencing (RNA-seq) to screen the downstream genes regulated by lactylation in PRRSV-infected cells, we found that PRRSV-induced lactylation activated the expression of heat shock 70 kDa protein 6 (HSPA6). Follow-up experiments showed that HSPA6 is important for PRRSV proliferation by negatively modulating interferon (IFN)-beta induction. Mechanistically, HSPA6 impeded the interaction between TNF-receptor-associated factor 3 (TRAF3) and inhibitor of nuclear factor kappa-B kinase subunit epsilon (IKK epsilon), thereby hindering the production of IFN-beta. Taken together, these results indicate that the activated lactate-lactylation-HSPA6 axis promotes viral growth by impairing IFN-beta induction, providing new therapeutic targets for the prevention and control of PRRSV infection. The results presented here also link lactylation to the virus life cycle, improving our understanding of epigenetic regulation in viral infection.IMPORTANCEAs a newly identified epigenetic modification, lactate-induced lactylation has received attentions because it plays important roles in gene expression and contributes to tumorigenesis and the innate immune response. Previous studies showed that many viruses upregulate cellular lactate levels; however, whether virus-elevated lactate induces lactylation and the subsequent biological significance of the modification to viral infection have not been reported. In this study, we demonstrated that porcine reproductive and respiratory syndrome virus (PRRSV) infection induced cellular lactylation, which, in turn, upregulated the expression of HSPA6, an IFN-negative regulator. We also dissected the mechanism by which HSPA6 negatively regulates IFN-beta production. To our knowledge, this is the first report to study virus-induced lactylation and establish the relationship between lactylation and virus infection. As a newly identified epigenetic modification, lactate-induced lactylation has received attentions because it plays important roles in gene expression and contributes to tumorigenesis and the innate immune response. Previous studies showed that many viruses upregulate cellular lactate levels; however, whether virus-elevated lactate induces lactylation and the subsequent biological significance of the modification to viral infection have not been reported. In this study, we demonstrated that porcine reproductive and respiratory syndrome virus (PRRSV) infection induced cellular lactylation, which, in turn, upregulated the expression of HSPA6, an IFN-negative regulator. We also dissected the mechanism by which HSPA6 negatively regulates IFN-beta production. To our knowledge, this is the first report to study virus-induced lactylation and establish the relationship between lactylation and virus infection.