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SARS-CoV-2 spike sequence dictates host TP53 activity and COVID-19 pathogenicity

ABSTRACT: SARS-CoV-2 induces widespread transcriptomic changes in host cells upon infection, in part through activation and modulation of innate immunity pathways and downstream gene regulation. However, the mechanisms by which SARS-CoV-2 and its evolutionary variants differentially affect host cell transcriptomic states remain largely unclear. Through chromatin proteomic (iDAPT-MS) analysis, we found that although SARS-CoV-2 and other pathogenic coronaviruses exhibit similar proteomic shifts on chromatin, SARS-CoV-2 uniquely promotes TP53 nuclear accumulation and activation. Parallel assessment of SARS-CoV-2 viral protein expression on host chromatin states (ATAC-seq) identifies intracellular spike protein as a key determinant of virus-mediated chromatin accessibility changes. Multilevel chromatin profiling reveals increased TP53 nuclear accumulation, TP53-associated chromatin accessibility changes, and TP53 target gene activation upon expression of SARS-CoV-2 alpha (B.1.1.7) and delta (B.1.617.2) spike variants relative to the ancestral spike sequence. TP53, ACE2, and furin cleavage are required for these changes, driving decreased cellular proliferation, increased cellular senescence, and increased cytokine release. Finally, BA.1 but not BA.2, BA.2.12.1, nor BA.4/BA.5 spike expression leads to attenuated TP53 activity and fusogenicity relative to ancestral spike. Our findings implicate spike-mediated host TP53 activation as a “rheostat” of COVID-19 pathogenicity.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Chlorocebus Sabaeus

SUBMITTER: Joao Paulo

LAB HEAD: Frank J. Slack

PROVIDER: PXD034648 | Pride | 2023-09-09

REPOSITORIES: Pride

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Project description:Rapid emergence of antigenic distinct SARS-CoV-2 variants implies a greater risk of reinfection as viruses can escape neutralizing antibodies induced by vaccination or previous viral exposure. Disease severity during COVID-19 depends on many variables such as age-related comorbidities, host immune status and genetic variation. The host immune response during infection with SARS-CoV-2 may contribute to disease severity, which can range from asymptomatic to severe with fatal outcome. Furthermore, the extent of host immune response activation may rely on underlying genetic predisposition for disease or protection. To address these questions, we performed immune profiling studies in mice with different genetic backgrounds - transgenic K18-hACE2 and wild-type 129S1 mice – subjected to reinfection with the severe disease-causing SARS-CoV-2 B.1.351 variant, 30 days after experimental milder BA.1 infection. BA.1 preinfection conferred protection against B.1.351-induced morbidity in K18-hACE2 mice but aggravated disease in 129S1 mice. We found that he cytokine/chemokine profile in B.1.351 re-infected 129S1mice is similar to that during severe SARS-CoV-2 infection in humans and is characterized by a much higher level of IL-10, IL-1β, IL-18 and IFN-γ , whereas in B.1.351 re-infected K18-hACE2 mice, the cytokine profile echoes the signature of naïve mice undergoing viral infection for the first time. Interestingly, the enhanced pathology observed in 129S1 mice upon reinfection cannot be attributed to a less efficient induction of adaptive immune responses to the initial BA.1 infection, as both K18-hACE2 and 129S1 mice exhibited similar B and T cell responses at 30 DPI against BA.1, with similar anti-BA.1 or B.1.351 spike-specific ELISA binding titers, levels of germinal center B-cells, and SARS-CoV-2-Spike specific tissue-resident T-cells. Long-term effects of BA.1 infection are associated with differential transcriptional changes in bronchoalveolar lavage-derived CD11c+ immune cells from K18-hACE2 and 129S1, with K18-hACE2 CD11c+ cells showing a strong antiviral defense gene expression profile whereas 129S1 CD11c+ cells showed a more pro-inflammatory response. In conclusion, initial infection with BA.1 induces cross-reactive adaptive immune responses in both K18-hACE2 and 129S1 mice, however the different disease outcome of reinfection seems to be driven by differential responses of CD11c+ cells in the alveolar space.

Project description:Memory B cells play a fundamental role in host defenses against viruses. This dataset aimed at understanding the recruitment and remodeling of the memory B cell repertoire in the context of BA.1-breakthrough infection in BNT162b2 mRNA vaccinated individuals. All four donors enrolled in the study had received a booster (3rd dose) of BNT162b2 mRNA vaccine and had no history of prior SARS-CoV-2 infection. All four experienced a documented breakthrough infection between 12/24/2021 and 01/30/2022 when BA.1 was responsible for > 85% of SARS-CoV-2 infections in France. All donors were sampled at Henri Mondor University Hospital (AP-HP, Paris France), and samples used for scRNA-seq were collected shortly after breakthrough infection (PO_M0 samples; between day 7 and day 18) and 5 to 6 months after infection (PO_M6 samples; between day 152 and day 173). Clinical and biological characteristics of these patients are summarized in the Patient_information.csv file. For each sample, an initial pool of 50.000 total peripheral CD3-CD14-CD56- CD19+ IgD- cells was always sorted and afterward, to enrich for cells of interest, only CD19+CD38low antibody secreting cells (ASCs), CD19hi IgD+ cells and SARS-CoV-2 Spike/RBD PE/tetramer positive B cells were sorted, leading to approximately 55000-60000 total sorted cells per sample. Sorted cells were then counted and up to 20 000 cells were loaded in the 10x Chromium Controller to generate single-cell gel-beads in emulsion. The scRNA-seq libraries for gene expression (mRNA), ADT and VDJ BCR libraries were generated using Chromium Next GEM Single Cell V(D)J Reagent Kit v.1.1 with Feature Barcoding (10x Genomics) according to the manufacturer’s protocol. PBMCs were initially isolated from venous blood samples via standard density gradient centrifugation and used after cryopreservation at -150°C. PBMCs were thawed using RPMI-1640 (Gibco) 10% FBS, washed twice and approximately 15x106 cells were then resuspended in 100µL PBS 2%FBS and incubated for 40 minutes at 4°C with a decoy tetramer (biotinylated Bovine Serum albumin coupled with BV785 streptavidin) and Hu-1 Spike, BA.1 Spike, Hu-1 RBD and BA.1 RBD tetramers (constructed using PE-labelled TotalSeqC® streptavidin with different barcodes for each individual antigens (see feature_reference.csv.gz files).). Cells were washed, resuspended in 100µL PBS 2%FBS and stained with a cocktail of fluorochrome conjugated (CD3, CD14 both APC-H7 at 1:100 each; CD15 and CD56 BV785 at 1:100 each, CD19 PECF594 at 1:100, IgD FITC at 1:100, CD38 PercP-Cy5.5 at 1:100) and TotalSeqC® (CD38, CD27, CD71, CD21, CD11c, CD39, FCRL5, CD95 all at 1:40 (all obtained from BioLegend)) antibodies for 40 minutes on ice. Viable cells were identified using a LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific, 1:200) incubated with conjugated antibodies. Two distinct sorts were performed for each donor: one at the early time-point (PO_M0) and one at the 6 months’ time-point (PO_M6).

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SARS-CoV-2 spike sequence dictates host TP53 activity and COVID-19 pathogenicity

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