Project description:This experiment aims to profile polyclonal antibody binding profiles in serum from vaccinated animals relative to antibody function in a virus neutralization assay. Rabbits received three vaccinations with a DNA vaccine encoding the spike protein of the SARS-CoV-2 index strain. Serum samples were selected based on a three-tier (low, intermediate, and high) capacity to cross-neutralize SARS-CoV-2 strains with known neutralization resistance. Following normalization of total anti-spike IgG levels, serum of each animal (n=3) were evaluated for antibody binding to 10mer cyclic constrained peptides spanning the entire spike protein and regions with known SARS-CoV-2 variant of concern spike mutations.
Project description:The experiment aims at characterizing the immune responses elicited by the BNT162b2 vaccine against SARS-CoV-2, initially administered in a two dose regimen (second dose after three weeks followinf the first dose) In particular the transcriptional landscape of circulating T and B lymphocytes has been profiled longitudinnaly by scRNA-seq coupleD with CITE-seq of 19 cell surface markers to better classify T cells subpopulations, LIBRA-seq to assess the Spike-specificity of BCRs and and V(D)J seq to also track T and B cell clones dynamics. Eeach sample was profiled before vaccination (T0), 21 days after the first dose (T1), 2 months after the first dose (1 month after the second dose) (T2). The immune responses were characterized using PBMC from 3 SARS-CoV-2 experienced donors (experiencing SARS-Cov-2 at least 4 months before the first vaccinatin) and 2 SARS-CoV-2 unexperienced donors.
Project description:COVID-19 remains a significant public health threat due to the ability of SARS-CoV-2 variants to evade the immune system and cause breakthrough infections. Although pathogenic coronaviruses such as SARS-CoV-2 and MERS-CoV lead to severe respiratory infections, how these viruses affect the chromatin proteomic composition upon infection remains largely uncharacterized. Here we used our recently developed integrative DNA And Protein Tagging (iDAPT) methodology to identify changes in host chromatin accessibility states and chromatin proteomic composition upon infection with pathogenic coronaviruses. SARS-CoV-2 infection induces TP53 stabilization on chromatin, which contributes to its host cytopathic effect. We mapped this TP53 stabilization to the SARS-CoV-2 spike and its propensity to form syncytia, a consequence of cell-cell fusion. Differences in SARS-CoV-2 spike variant-induced syncytia formation modify chromatin accessibility, cellular senescence, and inflammatory cytokine release via TP53. Our findings suggest that differences in syncytia formation alter senescence-associated inflammation, which varies among SARS-CoV-2 variants.
Project description:COVID-19 remains a significant public health threat due to the ability of SARS-CoV-2 variants to evade the immune system and cause breakthrough infections. Although pathogenic coronaviruses such as SARS-CoV-2 and MERS-CoV lead to severe respiratory infections, how these viruses affect the chromatin proteomic composition upon infection remains largely uncharacterized. Here we used our recently developed integrative DNA And Protein Tagging (iDAPT) methodology to identify changes in host chromatin accessibility states and chromatin proteomic composition upon infection with pathogenic coronaviruses. SARS-CoV-2 infection induces TP53 stabilization on chromatin, which contributes to its host cytopathic effect. We mapped this TP53 stabilization to the SARS-CoV-2 spike and its propensity to form syncytia, a consequence of cell-cell fusion. Differences in SARS-CoV-2 spike variant-induced syncytia formation modify chromatin accessibility, cellular senescence, and inflammatory cytokine release via TP53. Our findings suggest that differences in syncytia formation alter senescence-associated inflammation, which varies among SARS-CoV-2 variants.
Project description:We generated LNP-mRNA encoding B.1.1.529 SARS-CoV-2 spike, and intramuscularly administered it in a human IgG and IgK knock-in mouse. Single cell VDJ-seq unveiled the sequences of human monoclonal antibodies targeting the B.1.1.529 SARS-CoV-2 spike protein.
Project description:Intranasal vaccines can prime or recruit to the respiratory epithelium mucosal immune cells capable of preventing transmission of SARS-CoV-2. We found that a single intranasal dose of serotype 5-based adenoviral vectors expressing either the receptor binding domain (Ad5-RBD) or the complete ectodomain (Ad5-S) of the SARS-CoV-2 spike protein was effective in inducing i) secretory and serum anti-spike IgA and IgG, ii) robust SARS-CoV-2-neutralizing activity in the serum and in respiratory secretions, iii) rigorous spike-directed T helper 1 cell/cytotoxic T cell immunity, and iv) protection of wild-type mice from a challenge with the SARS-CoV-2 beta variant. Our data confirm and extend previous studies reporting promising preclinical results on vector-based intranasal SARS-CoV-2 vaccination, and support the potential of this approach to elicit mucosal immunity for preventing reinfection and transmission of SARS-CoV-2 more effectively than the currently available vaccines.
Project description:We investigated the kinetics, breadth, magnitude, and level of cross-reactivity of IgG antibodies against SARS-CoV-2 and heterologous seasonal (HCoV-NL63, -229E, -OC43 and -HKU1) and epidemic coronaviruses (SARS-CoV, hCoV-MERS) at the clonal level in patients with mild or severe COVID-19 as well as in disease control patients. We assessed IgG antibody reactivity to nucleocapsid and spike antigens using protein microarray. A cutoff was set at the average plus 3 times the SD of 20 nonreactive cultures with a minimum MFI of 1000.
Project description:We have developed a SARS-CoV-2 PsV system that expresses the spike protein on the viral surface and carries spike protein genomic sequences, enabling this pseudovirus to activate infection signaling pathways through ACE2 and stimulate intracellular spike protein gene expression. To understand the impact of SARS-CoV-2 on hematopoietic system, we infected heman HSPCs with Omicron PsV and analyzed changes in transcriptome
Project description:The viral RNA-dependent RNA polymerase (replicase) from Venezuelan equine encephalitis virus constitutes a vital component of the bipartite trans-amplifying mRNA vaccine. In this vaccine strategy aimed at targeting SARS-CoV-2, the replicase mRNA is administered alongside the mRNA encoding the SARS-CoV-2 spike protein. Our investigation sought to determine whether the replicase induces amplification of cellular mRNAs. To this end, cells were transfected with mRNAs encoding the replicase and SARS-CoV-2 spike protein, while control groups received transfections of mRNAs encoding an unrelated protein along with the SARS-CoV-2 spike. We observed no significant upregulation of genes in the treatment group compared to the control group. This suggests that the replicase does not induce off-target amplification of cellular mRNAs.
Project description:We here identified that the trimeric spike protein of SARS-CoV-2 could bind to TLR4 directly and robustly activate downstream signaling in monocytes and neutrophils. Moreover, specific TLR4 or NFKB inhibitor, or knockout of MyD88 could significantly block IL-1B induction by spike protein. We thus reveal that spike protein of SARS-CoV-2 functions as a potent stimulus causing TLR4 activation and sepsis related abnormal responses.