Cyclophilin Inhibitors Block Arterivirus Replication by Interfering with Viral RNA Synthesis

Abstract

e Virus replication strongly depends on cellular factors, in particular, on host proteins. Here we report that the replication of the arteriviruses equine arteritis virus (EAV) and porcine reproductive and respiratory syndrome virus (PRRSV) is strongly affected by low-micromolar concentrations of cyclosporine A (CsA), an inhibitor of members of the cyclophilin (Cyp) family. In infected cells, the expression of a green fluorescent protein (GFP) reporter gene inserted into the PRRSV genome was inhibited with a half-maximal inhibitory concentration (IC 50 ) of 5.2 M, whereas the GFP expression of an EAV-GFP reporter virus was inhibited with an IC 50 of 0.95 M. Debio-064, a CsA analog that lacks its undesirable immunosuppressive properties, inhibited EAV replication with an IC 50 that was 3-fold lower than that of CsA, whereas PRRSV-GFP replication was inhibited with an IC 50 similar to that of CsA. The addition of 4 M CsA after infection prevented viral RNA and protein synthesis in EAV-infected cells, and CsA treatment resulted in a 2.5-to 4-log-unit reduction of PRRSV or EAV infectious progeny. A complete block of EAV RNA synthesis was also observed in an in vitro assay using isolated viral replication structures. The small interfering RNA-mediated knockdown of Cyp family members revealed that EAV replication strongly depends on the expression of CypA but not CypB. Furthermore, upon fractionation of intracellular membranes in density gradients, CypA was found to cosediment with membranous EAV replication structures, which could be prevented by CsA treatment. This suggests that CypA is an essential component of the viral RNA-synthesizing machinery.

Suppression of Coronavirus Replication by Cyclophilin Inhibitors

“…also reported that EAV replication was inhibited by knockdown of CypA expression, but not by knockdown of CypB expression [33]. The titers of virus progeny decreased approximately fourfold in CypA-knockdown cells as compared to those in the non-targeted cells.…”

“…The used virus may have been one of the naturally CsA-resistant viruses that are sometimes present in the virus pool. Several other RNA viruses require CypA or CypB in the replication process, as has been described for HCV, equine arterivirus (EAV), Japanese encephalitis virus, VSV, and West Nile virus (WNV) [16,17,33,34,35]. Therefore, de Wilde et al .…”

“…recently reported that CsA inhibits arteriviruses, which are distantly related to CoVs; together with the Coronaviridae and Roniviridae families, they constitute the order Nidovirales [33]. CsA as an immunosuppressive agent and Debio-064 as a non-immunosuppressive agent inhibited EAV and PRRSV replication.…”

Cyclophilin A Binds to the Viral RNA and Replication Proteins, Resulting in Inhibition of Tombusviral Replicase Assembly

“…The list of these viruses includes flaviviruses and coronaviruses (5,71). HCV usurps the cytosolic CypA to facilitate the assembly of the HCV VRC through influence on the cleavage of the NS5A-NS5B fusion protein and the folding of the NS5A and NS5B RdRp proteins (59,(72)(73)(74).…”

Effects on pig immunophysiology, PBMC proteome and brain neurotransmitters caused by group mixing stress and human-animal relationship

“…Downregulated chaperones include HSP90B1 [56], peptidyl-prolyl cis-trans isomerase A (PPIA) [57–60] and chaperonin containing TCP1, subunit 2 (Beta) (CCT2) [61,62]. These proteins are downregulated at t2, possibly related to the positive action of GC in proteasome activity and protein turnover [63,64].…”