Cleavage of the selective autophagy receptor NBR1 by the PDCoV main protease NSP5 impairs autophagic degradation of the viral envelope protein

Porcine deltacoronavirus (PDCoV) is an emerging enteropathogenic coronavirus that causes severe diarrhea in neonatal piglets worldwide and presents a significant public health threat due to its potential for cross-species transmission. Selective macroautophagy/autophagy, mediated by autophagy receptors such as NBR1 (NBR1 autophagy cargo receptor), plays a key role in restricting viral infection and modulating the host immune response. In this study, we revealed that overexpression of NBR1 inhibits PDCoV replication, while its knockdown increases viral titers. Further analysis demonstrated that NBR1 interacts with the PDCoV envelope (E) protein independently of ubiquitination, directing it to phagophores for autophagic degradation to limit viral proliferation. To counteract this defense, PDCoV 3C-like protease, encoded by NSP5, cleaves porcine NBR1 at glutamine 353 (Q353), impairing its selective autophagy function and antiviral activity. Additionally, we demonstrated that NSP5 proteases from other coronaviruses including PEDV, TGEV, and SARS-CoV-2 also cleave NBR1 at the same site, suggesting that coronaviruses employ a conserved strategy of NSP5-mediated cleavage of NBR1 to evade host antiviral responses and facilitate infection. Overall, our study underscores the importance of NBR1-mediated selective autophagy in the host's defense against PDCoV and reveals a strategy by which PDCoV evades autophagic mechanisms to promote successful infection.<b>Abbreviation</b>: Cas9: CRISPR-associated protein 9; CC1: coiled-coil 1; Co-IP: co-immunoprecipitation; CRISPR: clustered regularly interspaced short palindromic repeats; GFP: green fluorescent protein; IFA: indirect immunofluorescence assay; KO: knockout; LIR: MAP1LC3/LC3-interacting region; mAb: monoclonal antibody; NBR1: NBR1 autophagy cargo receptor; NBR1-C: C-terminal fragment of NBR1; NBR1-N: N-terminal fragment of NBR1; OPTN: optineurin; pAb: polyclonal antibody; PB1: Phox/BEM1 domain; PDCoV: porcine deltacoronavirus; PEDV: porcine epidemic diarrhea virus; Q353A: a NBR1 construct with the glutamine (Q) residue at position 353 replaced with glutamic acid (A); SARS-CoV-2: severe acute respiratory syndrome coronavirus 2; SQSTM1: sequestosome 1; TCID₅₀: 50% tissue culture infective dose; TGEV: porcine transmissible gastroenteritis virus; UBA: ubiquitin-associated domain; Ub: ubiquitin; WT: wild type; ZZ: ZZ-type zinc finger domain.