The novel Nsp9-interacting host factor H2BE promotes PEDV replication by inhibiting endoplasmic reticulum stress-mediated apoptosis

Abstract

Porcine epidemic diarrhoea (PED) caused by porcine epidemic diarrhoea virus (PEDV) has led to significant economic losses in the swine industry worldwide. Histone Cluster 2, H2BE (HIST2H2BE), the main protein component in chromatin, has been proposed to play a key role in apoptosis. However, the relationship between H2BE and PEDV remains unclear. In this study, H2BE was shown to bind and interact with PEDV nonstructural protein 9 (Nsp9) via immunoprecipitation-mass spectrometry (IP-MS). Next, we verified the interaction of Nsp9 with H2BE by immunoprecipitation and immunofluorescence. H2BE colocalized with Nsp9 in the cytoplasm and nuclei. PEDV Nsp9 upregulated the expression of H2BE by inhibiting the expression of IRX1. We demonstrated that overexpression of H2BE significantly promoted PEDV replication, whereas knockdown of H2BE by small interfering RNA (siRNA) inhibited PEDV replication. Overexpression of H2BE led to significantly inhibited GRP78 expression, phosphorylated PERK (p-PERK), phosphorylated eIF2 (p-eIF2), phosphorylated IRE1 (p-IRE1), and phosphorylated JNK (p-JNK); negatively regulated CHOP and Bax expression and caspase-9 and caspase-3 cleavage; and promoted Bcl-2 production. Knocking down H2BE exerted the opposite effects. Furthermore, we found that after deletion of amino acids 1–28, H2BE did not promote PEDV replication. In conclusion, these studies revealed the mechanism by which H2BE is associated with ER stress-mediated apoptosis to regulate PEDV replication. Nsp9 upregulates H2BE. H2BE plays a role in inhibiting apoptosis and thus facilitating viral replication, which depends on the N-terminal region of H2BE (amino acids 1–28). These findings provide a reference for host–PEDV interactions and offer the possibility for developing strategies for PEDV decontamination and prevention.

PEDV inhibits HNRNPA3 expression by miR-218-5p to enhance cellular lipid accumulation and promote viral replication

“…The mouse polyclonal anti-PEDV N and anti-PEDV NSP9 antibodies were stocked in our laboratory ( 43 ). Rabbit polyclonal anti-HNRNPA3 antibody (25142-1-AP), mouse monoclonal anti-FASN antibody (66591-1-lg), rabbit polyclonal anti-SREBF1 antibody (14088–1-AP), mouse monoclonal anti-ACC antibody (67373–1-lg), and mouse monoclonal anti-β-actin (66009-1-lg) were from Proteintech.…”

Insights into the Activation of Unfolded Protein Response Mechanism during Coronavirus Infection

“…PEDV Nsp9 interacts with H2BE, causing H2BE overexpression by inhibiting IRX1, a regulator of H2BE expression. H2BE overexpression inhibits the PDEV-induced upregulation of PERK, IRE1, and their respective apoptotic executioners, thus suppressing cell death and contributing to viral replication [75]. PEDV nsp14, via its N7-TMase domain, can suppress Grp78 expression.…”

A Comprehensive View on the Protein Functions of Porcine Epidemic Diarrhea Virus

“…Promotes the replication of PEDV [36] Nsp3 PLpro, regulates the deubiquitination of RIG-I and STING, inhibits IFN-β and IFN-λ1 [37,38] Nsp4 Upregulates pro-inflammatory cytokines and chemokines expression (IL-1α, IL-1β, TNF-α, CCL2, CCL5, and CXCL8) [39] Nsp5 3C-like protease，IFN antagonist [40,41] Nsp7 Inhibits type I IFN [42,43] Nsp8 Inhibits type III IFN [22] Nsp9 Inhibits ERS-mediated apoptosis [44] Nsp10 Essential for viral replication, upregulates IL-2, IL-4, IL-10, TNF-α, and IFN-γ [45,46] Nsp12 RdRp, viral replication [47] Nsp13 HEL, inhibits bidirectional IgG transport by FcRn [48] Nsp14 ExoN, suppresses ER stress-induced GRP78, acts as NF-κB pathway antagonist, downregulates pro-inflammatory cytokines [49,50] Nsp15 EndoU, inhibits IFN-β and IRF3, downregulates CCL5, CXCL8, CXCL10, OAS, MXs, STAT1, and IRF9 [51,52] Nsp16 2 ′ -O-MTase, downregulates the activities of RIG-I/MDA5-mediated IFN-β and ISRE [53]…”

“…Furthermore, the nsp9 has the capacity to increase the expression of H2BE, thereby suppressing ERS-induced apoptosis and promoting viral replication. This process is reliant on the N-terminal domain of H2BE (amino acids 1-28) [44].…”