Zoonotic introduction of novel coronaviruses may encounter preexisting immunity in humans. Using diverse assays for antibodies recognizing SARS-CoV-2 proteins, we detect preexisting humoral immunity. SARS-CoV-2 spike glycoprotein (S)-reactive antibodies were detectable by a flow cytometry-based method in SARS-CoV-2-uninfected individuals and were particularly prevalent in children and adolescents. They were predominantly of the IgG class and targeted the S2 subunit. By contrast, SARS-CoV-2 infection induced higher titers of SARS-CoV-2 S-reactive IgG antibodies, targeting both the S1 and S2 subunits, and concomitant IgM and IgA antibodies, lasting throughout the observation period. Notably, SARS-CoV-2-uninfected donor sera exhibited specific neutralizing activity against SARS-CoV-2 and SARS-CoV-2 S pseudotypes. Distinguishing preexisting and de novo immunity will be critical for our understanding of susceptibility to and the natural course of SARS-CoV-2 infection.

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Several related human coronaviruses (HCoVs) are endemic in the human population, causing mild respiratory infections 1 . Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the etiologic agent of Coronavirus disease 2019 , is a recent zoonotic infection that has quickly reached pandemic spread 2,3 . Zoonotic introduction of novel coronaviruses is thought to occur in the absence of pre-existing immunity in the target human population. Using diverse assays for detection of antibodies reactive with the SARS-CoV-2 Spike (S) glycoprotein, we demonstrate the presence of pre-existing immunity in uninfected and unexposed humans to the new coronavirus. SARS-CoV-2 S-reactive antibodies, exclusively of the IgG class, were readily detectable by a sensitive flow cytometry-based method in SARS-CoV-2-uninfected individuals with recent HCoV infection and targeted the S2 subunit. In contrast, SARS-CoV-2 infection induced higher titres of SARS-CoV-2 Sreactive IgG antibodies, as well as concomitant IgM and IgA antibodies throughout the observation period of 6 weeks since symptoms onset. HCoV patient sera also variably reacted with SARS-CoV-2 S and nucleocapsid (N), but not with the S1 subunit or the receptor binding domain (RBD) of S on standard enzyme immunoassays. Notably, HCoV patient sera exhibited specific neutralising activity against SARS-CoV-2 S pseudotypes, according to levels of SARS-CoV-2 S-binding IgG and with efficiencies comparable to those of COVID-19 patient sera. Distinguishing pre-existing and de novo antibody responses to SARS-CoV-2 will be critical for serology, seroprevalence and vaccine studies, as well as for our understanding of susceptibility to and natural course of SARS-CoV-2 infection. ResultsImmune cross-reactivity among seasonally spreading human coronaviruses (HCoVs) has long been hypothesised to provide cross-protection, albeit transient, against infection with distinct HCoV types 1,4,5 . To determine the degree of cross-reactivity between HCoVs and the recently introduced zoonotic coronavirus SARS-CoV-2, we developed a sensitive flow cytometry-based assay for detection of SARS-CoV-2-binding antibodies. Sera from COVID-19 patients at University College London Hospitals (UCLH) (Table S1), contained high levels of IgG, IgM and IgA antibodies recognising the wild-type Spike (S) glycoprotein of SARS-CoV-2 expressed on the surface of HEK293T cells, whereas control sera did not (Fig. 1a). Notably, sera from a proportion patients with confirmed HCoV infection collected before or during the early spread of SARS-CoV-2 in the UK (Table S1), also contained SARS-CoV-2 S-specific antibodies (Fig. 1a). However, the latter sera contained only lower levels of S-specific IgG and no IgM or IgA antibodies, which clearly distinguished them from COVID-19 patient sera (Fig. 1a). The SARS-CoV-2 S protein is proteolytically processed into the S1 and S2 subunits that mediate target cell attachment and entry, respectively 6,7 . S2 exhibits a higher degree of homology among coronaviruses than S1 (Extended data Fig. 1...

The majority of currently circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viruses have mutant spike glycoproteins that contain the D614G substitution. Several studies have suggested that spikes with this substitution are associated with higher virus infectivity. We use cryo-electron microscopy to compare G614 and D614 spikes and show that the G614 mutant spike adopts a range of more open conformations that may facilitate binding to the SARS-CoV-2 receptor, ACE2, and the subsequent structural rearrangements required for viral membrane fusion.

Highly polymorphic major histocompatibility complex (MHC) molecules are at the heart of adaptive immune responses, playing crucial roles in many kinds of disease and in vaccination. We report that breadth of peptide presentation and level of cell surface expression of class I molecules are inversely correlated in both chickens and humans. This relationship correlates with protective responses against infectious pathogens including Marek's disease virus leading to lethal tumours in chickens and human immunodeficiency virus infection progressing to AIDS in humans. We propose that differences in peptide binding repertoire define two groups of MHC class I molecules strategically evolved as generalists and specialists for different modes of pathogen resistance. We suggest that differences in cell surface expression level ensure the development of optimal peripheral T cell responses. The inverse relationship of peptide repertoire and expression is evidently a fundamental property of MHC molecules, with ramifications extending beyond immunology and medicine to evolutionary biology and conservation.DOI: http://dx.doi.org/10.7554/eLife.05345.001

SummaryClathrin-coated vesicles form by rapid assembly of discrete coat constituents into a cargo-sorting lattice. How the sequential phases of coat construction are choreographed is unclear, but transient protein-protein interactions mediated by short interaction motifs are pivotal. We show that arrayed Asp-Pro-Phe (DPF) motifs within the early-arriving endocytic pioneers Eps15/R are differentially decoded by other endocytic pioneers Fcho1/2 and AP-2. The structure of an Eps15/R⋅Fcho1 μ-homology domain complex reveals a spacing-dependent DPF triad, bound in a mechanistically distinct way from the mode of single DPF binding to AP-2. Using cells lacking FCHO1/2 and with Eps15 sequestered from the plasma membrane, we establish that without these two endocytic pioneers, AP-2 assemblies are fleeting and endocytosis stalls. Thus, distinct DPF-based codes within the unstructured Eps15/R C terminus direct the assembly of temporary Fcho1/2⋅Eps15/R⋅AP-2 ternary complexes to facilitate conformational activation of AP-2 by the Fcho1/2 interdomain linker to promote AP-2 cargo engagement.

The coronaviral spike is the dominant viral antigen and the target of neutralizing antibodies. We show that SARS-CoV-2 spike binds biliverdin and bilirubin, the tetrapyrrole products of heme metabolism, with nanomolar affinity. Using cryo–electron microscopy and x-ray crystallography, we mapped the tetrapyrrole interaction pocket to a deep cleft on the spike N-terminal domain (NTD). At physiological concentrations, biliverdin significantly dampened the reactivity of SARS-CoV-2 spike with immune sera and inhibited a subset of neutralizing antibodies. Access to the tetrapyrrole-sensitive epitope is gated by a flexible loop on the distal face of the NTD. Accompanied by profound conformational changes in the NTD, antibody binding requires relocation of the gating loop, which folds into the cleft vacated by the metabolite. Our results indicate that SARS-CoV-2 spike NTD harbors a dominant epitope, access to which can be controlled by an allosteric mechanism that is regulated through recruitment of a metabolite.