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Comparative Proteomic Profiling of Variants SARS-CoV-2 Virus-Like-Particles (VLPs) Incubated Cells

ABSTRACT: The COVID-19 pandemic is arguably the most important global threat to humankind that the world has seen in the 21st century. Since the initial outbreak of SARS-CoV-2 in 2019, the virus genome is continuously mutating that may affect virus characteristics such as infectivity, transmission, and pathogenicity. We recently found that two consecutive amino acid mutations R203K and G204R in the nucleocapsid (N) protein, are linked with higher viral load and severity in COVID-19 patients. Here, we performed comparative total proteomic analysis of HEK293T (stably expressing ACE2/TMPRSS2) cell lines infected with wildtype (WT) VLP, KR-mutant (KR) VLP with two mutations R203K/G204R in the nucleocapsid protein, and two additional controls D614G-KR (containing both R203K/G204R and spike D614G mutations) and D614G (only spike D614G mutation). Our data reveal enhanced expression of immune response proteins in KR VLP infected cells.

INSTRUMENT(S): timsTOF Pro 2

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Kidney

DISEASE(S): Disease Free

SUBMITTER: Muhammad Shuaib

LAB HEAD: Muhammad Shuaib

PROVIDER: PXD034557 | Pride | 2023-04-15

REPOSITORIES: Pride

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Project description:Human papilloma (HPV) virus-like particle (VLP) vaccines were recently licensed. Though neutralizing antibody titers are thought to be the main effectors of protection against infection, early predictors of long-term efficacy are not yet defined and a comprehensive understanding of innate and adaptive immune responses to vaccination is still lacking. Here, microarrays were used to compare the gene expression signature in HPV-16 L1 VLP-stimulated PBMC from 17 vaccine and 4 placebo recipients before vaccination, and 1 month after receiving the second immunization. Vaccination with HPV-16 L1 VLP was associated with modulation of genes involved in the inflammatory/defense response, cytokine, interferon and cell cycle pathways in VLP-stimulated PBMC. In addition, there was up-regulation of probesets associated with cytotoxic (GZMB, TNFSF10) and regulatory (INDO, CTLA4) activities. The strongest correlations with neutralizing antibody titers were found for cyclin d2 (CCND2) and galectin (LGALS2). Twenty-two differentially expressed probesets were selected for confirmation by RT-PCR in an independent sample set. Agreement with the microarray data was seen for over two-thirds of these probesets. Up-regulation of immune/defense response genes by VLP, in particular interferon-induced genes was observed in PBMC collected prior to vaccination, with many of these genes being further induced following vaccination. In conclusion, we used gene expression profiling to identify important innate and adaptive response related- genes induced by vaccination with HPV VLP. Further studies are needed to identify gene expression signatures of immunogenicity and long-term protection with potential utility in prediction of long-term HPV vaccination outcomes in clinical trials. Experiment Overall Design: Microarrays (Affymetrix Human Focus) were used to compare the gene expression signature in PBMCs stimulated for 3 days with media alone, Sf9/baculovirus insect cell lysate, and HPV-16 L1 VLP expressed from baculovirus-infected Sf9 insect cells from 17 vaccine and 4 placebo recipients before vaccination, and 1 month after receiving the second immunization (2 months after the initial immunization. Post-vaccination baculovirus sample for Pt078 and pre-vaccination baculovirus sample for Pt082 failed QC.

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Comparative Proteomic Profiling of Variants SARS-CoV-2 Virus-Like-Particles (VLPs) Incubated Cells

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