Difference in receptor usage between SARS coronavirus and SARS-like coronavirus of bat origin

State key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China; Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing, China; CSIRO Livestock Industries, Australian Animal Health Laboratory and Australian Biosecurity Cooperative Research Center for Emerging Infectious Diseases, Geelong, Australia; and School of Life Science, East China Normal University, Shanghai, China

^* To whom correspondence should be addressed. Email: Linfa.wang{at}csiro.au. zlshi{at}wh.iov.cn.

	Abstract

Severe acute respiratory syndrome (SARS) was caused by the SARS-associated coronavirus (SARS-CoV), which uses the angiotensin converting enzyme 2 (ACE2) as its receptor for cell entry. A group of SARS-like coronaviruses (SL-CoVs) has been identified in horseshoe bats. SL-CoVs and SARS-CoVs share an identical genome organization and high sequence identities with the main exception of the N-terminus of the spike protein (S), known to be responsible for receptor binding in coronaviruses. In this study, we investigated the receptor usage of the SL-CoV S by combining an HIV-based pseudovirus system with cell lines expressing the ACE2 molecules of human, civet or horseshoe bat. In addition to full-length S of SL-CoV and SARS-CoV, a series of S chimeras was constructed by inserting different sequences of the SARS-CoV S into the SL-CoV S backbone. Several important observations were made from this study. First, the SL-CoV S was unable to use any of the three ACE2 molecules as its receptor. Second, the SARS-CoV S failed to enter cells expressing the bat ACE2. Third, the chimeric S covering the previously defined receptor-binding domain gained their ability to enter cells via human ACE2, albeit with different efficiency for different constructs. Fourth, a minimal insert region (aa 310-518) was found to be sufficient to convert the SL-CoV S from ACE2-non-binding to human ACE2-binding, indicating that the SL-CoV S is largely compatible to SARS-CoV S protein both in structure and function. The significance of these findings in relation to virus origin, virus recombination and host-switching is discussed.