Mechanistic study of PEDV NSP3-mediated regulation of host innate immune responses via ubiquitin removal and UPR pathway modulation

The endoplasmic reticulum (ER) maintains protein-folding homeostasis and initiates Antiviral responses via the unfolded protein response (UPR). Porcine epidemic diarrhea virus (PEDV) has evolved to exploit this process through its nonstructural protein 3 (NSP3). Here, we identify NSP3 as a multifunctional Deubiquitinase that remodels ER stress signaling and redox homeostasis to facilitate viral replication and immune evasion. NSP3 suppresses GRP78 expression while selectively activating PERK-eIF2α-ATF4 and IRE1α-XBP1s pathways but inhibits ATF6 nuclear translocation. Mechanistically, NSP3's PLP2 domain removes K48- and K63-linked ubiquitin chains from KEAP1, IRE1α, and ATF6, thereby fine-tuning their activity. Deubiquitination of KEAP1 promotes KEAP1-CUL3 complex stability and enhances NRF2 degradation, disrupting the antioxidant axis (NRF2-NQO1-HMOX1) while activating the ATF4-ERO1A-NOX4 pathway to elevate Reactive Oxygen Species (ROS) levels without cytotoxicity. Functionally, activation of the PERK-NRF2 axis suppresses PEDV replication, whereas inhibition of either component enhances viral growth, highlighting a cooperative Antiviral role. In contrast, activation of the IRE1α-XBP1s branch promotes replication but can be antagonized by NRF2 activation, indicating crosstalk between these pathways. Although ATF6 alone has minimal impact, its modulation by NRF2 shapes viral replication outcomes. Collectively, our findings reveal that PEDV NSP3 orchestrates differential deubiquitination of ER stress sensors to manipulate UPR-ROS signaling and redox balance, thereby subverting host defenses and promoting viral survival, providing mechanistic insight into coronavirus-host interactions and potential Antiviral targets.

Immune evasion; NSP3; Oxidative stress; PEDV; Unfolded protein response.