Initial blots for Figures 5D,E,G,H,J are provided in the Supplementary Material. Classical Swine Fever Computer virus Contamination Induces Endoplasmic Reticulum Stress-Induced Autophagy in Cultured Porcine Cell Lines ER stress-mediated autophagy Rabbit Polyclonal to GNAT2 has been well studied due to its significant functions in many diseases (Rashid et al., 2015; Senft and Ronai, 2015; Cybulsky, 2017). contamination activates the PERK and IRE1 pathways and autophagy in cultured porcine kidney cell lines (PK-15) and macrophage cell lines (3D4/2), and pharmacological regulation of ER stress remarkably changed autophagic activities induced by CSFV, suggesting that CSFV-induced autophagy can GSK-3326595 (EPZ015938) be mediated by ER stress possibly the PERK and IRE1 pathway. Furthermore, treatment with ER stress regulators significantly altered copy numbers of genes, expression of Npro proteins, and viral titers in CSFV-infected cells or in cells treated with autophagy regulators prior to CSFV infection, suggesting the requirement of ER stress-mediated autophagy for CSFV replication and genus in the family, CSFV contains an approximate 12.3-kb single-stranded sense RNA (ss(+)RNA) genome, encoding four structural proteins (C, Erns, E1, and E2) and eight non-structural proteins (Npro, P7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B) (Tautz et al., 1999; Lefkowitz et al., 2018). Acute contamination of CSFV causes persistent high fever, hemorrhages in multiple organs, and neurological, respiratory, and gastrointestinal symptoms in pigs with a very high mortality rate, leading to huge economic losses to the pig industry worldwide (Moennig, 1992; Lohse et al., 2012). Although clinical vaccinations can effectively prevent the outbreaks of CSF, there are no therapeutic drugs currently available on the market. CSFV replicates in leukocytes, especially in mononuclear macrophages, causing structural injuries and functional disorders in immune organs and subsequent immunosuppression in the diseased pigs (Floegel-Niesmann et al., 2003; Ji et al., 2015). Many viruses have evolutionarily developed specific or nonspecific strategies to evade host immune responses, during which many biological processes mediating the interplay between computer virus and host can be used for maintaining effective replication, GSK-3326595 (EPZ015938) contamination, and pathogenesis of viruses (Pei et al., 2014; Peacock et al., 2017; Ying et al., 2018). However, mechanisms involved in CSFV replication and pathogenesis still need further investigations. Endoplasmic reticulum (ER) is an important membranous organelle in eukaryotic cells. Homeostasis of the ER is usually a guarantee of maintaining normal cell activities. When the cells are exposed to stimuli including hypoxia, calcium overload, and free radical attack, dysfunctions of the ER occur and lead to accumulation of the misfolded/unfolded proteins in the lumen of ER and the imbalance of calcium homeostasis, resulting in ER stress (To Sing et al., 2015; Cybulsky, 2017). The ER responds to the burden of ER stress by activating a set of intracellular signaling pathways, known as the unfolded protein response (UPR), to restore normal function of the ER. There are three branches of the UPR: protein kinase R-like ER kinase (PERK), inositol requiring enzyme 1 (IRE1), and activating transcription factor-6 (ATF-6), which are not independent GSK-3326595 (EPZ015938) and together constitute GSK-3326595 (EPZ015938) a complex signaling network (Ron and Walter, 2007; Cybulsky, 2017). Under normal physiological conditions, glucose-regulated protein 78 (GRP78) binds to the three sensor proteins and inhibits their activities. Upon ER stress, GRP78 dissociates from the sensors and binds to the unfolded/misfolded proteins, and the released sensors are activated and initiate the following signaling pathways to alleviate ER stress through increasing protein-folding capacity of the ER, inhibiting global protein synthesis, and enhancing the ER-associated protein degradation (ERAD) of misfolded/unfolded proteins (Cybulsky, 2017). All these reactions aim to relieve the burden of ER and if they fail, they can trigger cellular dysfunction and eventually lead to cell death (Forman et al., 2003). In the past few years, the essential roles of the UPR have been implicated in many mammalian diseases, especially in viral diseases. Induction of ER stress and activation of the UPR signaling are general host responses to flavivirus and coronavirus infection, because replication of these viruses is closely associated with ER-derived membranes, and large amounts of viral proteins inevitably disturb the ER homeostasis and cause ER stress (Lin et al., 2002; Ambrose and Mackenzie, 2011; Pe?a and Harris, 2011; Fung and Liu, 2014). Our previous studies have revealed that CSFV infection induces the ER stress in the cultured porcine kidney PK-15 cells, benefiting its replication by activating the IRE1 pathway (He et al., 2017). However, the relationship between CSFV infection and ER stress-driven UPR, and the underlying mechanisms remain unclear. Autophagy is an evolutionarily conserved cellular degradation and recycling process in eukaryotic cells, which contains three major types: microautophagy, macroautophagy (hereafter referred to as autophagy), and chaperone-mediated autophagy. Of these three types, autophagy is the best studied autophagic process (Deretic and Klionsky, 2018). During autophagic process, cytoplasmic components are sequestered into and infection of CSFV still needs to be clarified. In the present study, basing on the established pig model for CSFV infection experiments further confirmed that CSFV-induced autophagy can be mediated by ER stress, and ER stress-mediated autophagy functions as a replication.