Insight into the evolution of nidovirus endoribonuclease based on the finding that nsp15 from porcine Deltacoronavirus functions as a dimer

Abstract

Nidovirus endoribonucleases (NendoUs) include nonstructural protein 15 (nsp15) from coronaviruses and nsp11 from arteriviruses, both of which have been reported to participate in the viral replication process and in the evasion of the host immune system. Results from a previous study of coronaviruses SARS-CoV, HCoV-229E, and MHV nsp15 indicate that it mainly forms a functional hexamer, whereas nsp11 from the arterivirus PRRSV is a dimer. Here, we found that porcine (PDCoV) nsp15 primarily exists as dimers and monomers Biological experiments reveal that a PDCoV nsp15 mutant lacking the first 27 amino acids of the N-terminal domain (Asn-1-Asn-27) forms more monomers and displays decreased enzymatic activity, indicating that this region is important for its dimerization. Moreover, multiple sequence alignments and three-dimensional structural analysis indicated that the C-terminal region (His-251-Val-261) of PDCoV nsp15 is 10 amino acids shorter and forms a shorter loop than that formed by the equivalent sequence (Gln-259-Phe-279) of SARS-CoV nsp15. This result may explain why PDCoV nsp15 failed to form hexamers. We speculate that NendoUs may have originated from XendoU endoribonucleases (XendoUs) forming monomers in eukaryotic cells, that NendoU from arterivirus gained the ability to form dimers, and that the coronavirus variants then evolved the capacity to assemble into hexamers. We further propose that PDCoV nsp15 may be an intermediate in this evolutionary process. Our findings provide a theoretical basis for improving our understanding of NendoU evolution and offer useful clues for designing drugs and vaccines against nidoviruses.

Development and utilization of an infectious clone for porcine deltacoronavirus strain USA/IL/2014/026

“…The function of the coronaviral endoribonuclease has been shown to be important for evading multiple host dsRNA sensors, including melanoma differentiation-associated protein 5 (MDA5), protein kinase R (PKR) and 2′-5′-oligoadenylate synthetase (OAS) [reviewed in ( Deng and Baker, 2018 )]. Sequence alignment from all four CoV genera and structural analysis showed that δ-CoV EnU contains the key residues that are required for endoribonuclease activity ( Deng and Baker, 2018 ; Zheng et al, 2018 ). As shown for EndoUs of mouse hepatitis virus, human coronavirus 229E, SARS-CoV-2, and PEDV, PDCoV EndoU may also interfere with host innate immunity ( Deng et al, 2017 , 2019 ; Kindler et al, 2017 ; Yuen et al, 2020 ).…”

Cryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics

“…Each Nsp15 protomer is composed of three domains including an N-terminal domain (ND) that is important for oligomerization, a variable middle domain (MD), and a C-terminal endonuclease (endoU) domain that shares homology with other endoU enzymes (Fig. 1a ) 17 . Nsp15 is only active as a hexamer, but the molecular requirement for Nsp15 oligomerization is unknown.…”

Porcine deltacoronavirus nsp15 antagonizes interferon-β production independently of its endoribonuclease activity

“…These results indicated that PDCoV nsp15 has evolved a novel mechanism for the antagonism of IFN production in an endoribonuclease activity-independent manner, distinct from that reported previously for the inhibition of IFN-β production by other CoV nsp15 s or arterivirus nsp11 s. The reason that results in these differences remains unclear. In terms of structure, the active form of endoribonuclease from PDCoV nsp15 is different from that of other CoV nsp15 s. PDCoV nsp15 is a mixture of dimers and monomers, while the nsp15 s of SARS-CoV, MHV, HCoV-229E, as well as MERS-CoV, function as a hexamer (Ricagno et al, 2006;Xu et al, 2006;Zhang et al, 2018;Zheng et al, 2018). Whether these structure differences result in different mechanisms used to antagonize IFN production remains further study.…”

“…Previous study has demonstrated that PDCoV nsp15 His219 (H219), H234, and Lys269 (K269) are the critical sites for its endoribonuclease activity (Zheng et al, 2018). To investigate whether or not PDCoV nsp15 mutants lacking endoribonuclease activity possess the ability to antagonize IFN-β production.…”