Research Article: The papain-like protease determines a virulence trait that varies among members of the SARS-coronavirus species

Date Published: September 24, 2018

Publisher: Public Library of Science

Author(s): Daniela Niemeyer, Kirstin Mösbauer, Eva M. Klein, Andrea Sieberg, Robert C. Mettelman, Anna M. Mielech, Ronald Dijkman, Susan C. Baker, Christian Drosten, Marcel A. Müller, Leo Poon.


SARS-coronavirus (CoV) is a zoonotic agent derived from rhinolophid bats, in which a plethora of SARS-related, conspecific viral lineages exist. Whereas the variability of virulence among reservoir-borne viruses is unknown, it is generally assumed that the emergence of epidemic viruses from animal reservoirs requires human adaptation. To understand the influence of a viral factor in relation to interspecies spillover, we studied the papain-like protease (PLP) of SARS-CoV. This key enzyme drives the early stages of infection as it cleaves the viral polyprotein, deubiquitinates viral and cellular proteins, and antagonizes the interferon (IFN) response. We identified a bat SARS-CoV PLP, which shared 86% amino acid identity with SARS-CoV PLP, and used reverse genetics to insert it into the SARS-CoV genome. The resulting virus replicated like SARS-CoV in Vero cells but was suppressed in IFN competent MA-104 (3.7-fold), Calu-3 (2.6-fold) and human airway epithelial cells (10.3-fold). Using ectopically-expressed PLP variants as well as full SARS-CoV infectious clones chimerized for PLP, we found that a protease-independent, anti-IFN function exists in SARS-CoV, but not in a SARS-related, bat-borne virus. This PLP-mediated anti-IFN difference was seen in primate, human as well as bat cells, thus independent of the host context. The results of this study revealed that coronavirus PLP confers a variable virulence trait among members of the species SARS-CoV, and that a SARS-CoV lineage with virulent PLPs may have pre-existed in the reservoir before onset of the epidemic.

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The coronaviruses (CoV, family Coronaviridae) are among the most relevant groups of viruses with zoonotic potential. CoVs are large, positive-sense, single-stranded RNA viruses that cause acute and prolonged infections in a variety of mammals and birds. Pathogenic human CoVs include members of the genus Alphacoronavirus, termed human coronavirus (HCoV)-NL63 and HCoV-229E, as well as members of the genus Betacoronavirus, termed HCoV-OC43 and HCoV-HKU1. These endemic viruses cause upper and lower respiratory tract infections in humans worldwide. Past zoonotic descent can be inferred for HCoV-OC43 and -229E, respectively [1–4]. Actual zoonotic acquisition is known for two betacoronaviruses that both cause severe lung disease in humans. The Middle East respiratory syndrome coronavirus (MERS-CoV) is a zoonotic agent that is frequently and repeatedly acquired by humans upon contact with dromedary camels in the Arabian Peninsula and parts of Africa [5, 6]. This virus seems to cause only limited human-to-human transmission, but is considered a major threat to global public health due to recurring nosocomial outbreaks that may facilitate onward adaptation to humans [7–9]. The severe acute respiratory syndrome (SARS)-CoV caused an epidemic with sustained human-to-human transmission during 2002 to 2003 in China and other countries, involving more than 8,000 notified infections with a case fatality proportion of about 10% [10, 11].

In an earlier study we have described SARS-related CoVs in European (Bulgarian) bat species belonging to the genus Rhinolophus [44]. Fig 1A shows a phylogeny of SARS-related beta-CoVs based on the PLP gene (981 bp fragment, genome position 4885 to 5829 in GenBank accession number AY310120). Based on standing classification criteria, the European bat-derived CoVs are conspecific with human SARS-CoV and in sister relationship to all Asian SARS-related CoVs.

SARS-CoV variants exist across Europe in widespread Rhinolophus bat species. It is important to understand whether these viruses constitute a risk for human infection [57]. Host tropism is mainly thought to be determined by the spike protein, but studies have shown that CoV populations in natural reservoirs can contain a plethora of spike variants, including variants that can directly mediate entry into human cells [16, 20]. Because of widespread recombination, spike proteins can be exchanged between viral genetic lineages in the reservoir. The spike protein may not sufficiently represent the variability of virulence traits in the reservoir.