Contents lists available at ScienceDirect Virology journal homepage: www.elsevier.com/locate/virology An “Old” protein with a new story: Coronavirus endoribonuclease is important for evading host antiviral defenses Xufang Deng⁎, Susan C. Baker Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University Chicago, Maywood, IL 60153, USA A R T I C L E I N F O Keywords: Coronavirus Nsp15 Endoribonuclease Double-stranded RNA Interferon Host recognition Antiviral defense A B S T R A C T Here we review the evolving story of the coronavirus endoribonuclease (EndoU). Coronavirus EndoU is encoded within the sequence of nonstructural protein (nsp) 15, which was initially identified as a component of the viral replication complex. Biochemical and structural studies revealed the enzymatic nature of nsp15/EndoU, which was postulated to be essential for the unique replication cycle of viruses in the order Nidovirales. However, the role of nsp15 in coronavirus replication was enigmatic as EndoU-deficient coronaviruses were viable and re- plicated to near wild-type virus levels in fibroblast cells. A breakthrough in our understanding of the role of EndoU was revealed in recent studies, which showed that EndoU mediates the evasion of viral double-stranded RNA recognition by host sensors in macrophages. This new discovery of nsp15/EndoU function leads to new opportunities for investigating how a viral EndoU contributes to pathogenesis and exploiting this enzyme for therapeutics and vaccine design against pathogenic coronaviruses. 1. History of coronavirus endoribonuclease We provide a brief outline of the major research findings related to coronavirus (CoV) endoribonucleases (EndoU) in Table 1. In the text below, we describe the experimental approaches that led to these findings and compare the activity of CoV EndoU with reports of other viral and host ribonucleases. 1.1. Identifying nsp15 as a component of the CoV replicase polyprotein Initial studies focused on identifying the products of the CoV re- plicase polyprotein, pp1ab (depicted in Fig. 1A). Heusipp et al. used a murine monoclonal antibody (10G11) that recognizes amino acid re- sidues 6158–6164 of human CoV 229E pp1ab (Heusipp et al., 1997). They identified a 41-kDa polypeptide in virus-infected cells and found that this polypeptide was a product of the viral 3C-like protease- mediated cleavage of pp1ab. This protein localizes to the perinuclear region, as detected by immunofluorescence assay, similar to other pp1ab-derived polypeptides (Heusipp et al., 1997). Shi and coworkers obtained similar findings while studying mouse hepatitis virus (MHV) (Shi et al., 1999). They generated a rabbit antiserum against amino acids 6679–6821 of MHV pp1ab and found that this antiserum detected a 35-kDa product in infected cells. They also found that this protein co- localized with de novo synthesized viral RNA, and therefore postulated that this viral protein associated with the viral RNA replication/ transcription machinery (Shi et al., 1999). Later, the corresponding polypeptides that these antibodies recognized were defined as the 15th cleavage product of pp1ab (called nsp15), counting from the amino- terminus to the carboxyl terminus of pp1ab (Ziebuhr et al., 2000; Snijder et al., 2003). 1.2. Bioinformatic analysis of nidovirus replicase polyproteins After the outbreak of Severe Acute Respiratory Syndrome (SARS) in 2002–2003, and once a CoV had been confirmed as the etiological agent of SARS, researchers intensively scrutinized CoV genomic se- quences to better understand this novel human pathogen. By com- parative genomic characterization of CoV replicases, Snijder et al. re- ported that the C-terminus of nsp15 has high sequence similarity to the Xenopus laevis poly(U)-specific endoribonuclease and therefore pre- dicted that nsp15 possesses EndoU activity (Snijder et al., 2003). Based on the available sequence information of viruses from the order Nido- virales at that time, the EndoU was considered a nidovirus-specific marker (called NendoU) (Fig. 1B) (Snijder et al., 2003). The members of the family Arteriviridae, including equine arteritis virus (EAV) and porcine respiratory and reproductive syndrome virus (PRRSV), also have EndoU domains within nsp11. However, it was later discovered that the presence of the EndoU domain is not universal in all nido- viruses. Nam Dinh virus, the first insect nidovirus belonging to the fa- mily Mesoniviridae, and roniviruses that infect invertebrates, do not https://doi.org/10.1016/j.virol.2017.12.024 Received 4 October 2017; Received in revised form 21 December 2017; Accepted 22 December 2017 ⁎ Corresponding author. E-mail address: