Investigating transcriptomic markers of SARS-CoV-2 response

Darmalinggam, Sathya (2025): Investigating transcriptomic markers of SARS-CoV-2 response: elucidating the role of Otoferlin as an interferon-stimulated gene. Dissertation, LMU München: Medizinische Fakultät

Abstract

With more than 700 million Severe Acute Respiratory Syndrome 2 (SARS-CoV-2) infections worldwide, understanding the host immune response upon infection is crucial in informing about and preventing a severe disease course, in this case Coronavirus-19 (COVID-19). Majority of studies undertaken thus far utilise adult disease cohorts and typically focus on antibody responses (natural infection or vaccine-induced), host antiviral mechanisms and viral evasion strategies. Questions such as the influence of age on the host immune response and identification of novel SARS-CoV-2 specific biomarkers still remain. To that end, we performed a comprehensive transcriptomic analysis on whole blood from a cohort comprising probands aged 2 weeks to 40 years old followed by functional experiments on elucidating the role of Otoferlin (OTOF) in the context of viral infections. In our transcriptomic analysis, by controlling for the effects of age, 246 genes were found to be significantly differentially expressed in the COVID cohort compared to healthy controls, including OTOF. When taking into account the effects of age, 4 genes (MMP8, OAS1, OAS2 and LY6E) were identified that showed strong differences in expression values in our COVID cohort compared to healthy controls. As much is unknown about the role of OTOF in the context of viral infections, we further conducted isoform analysis, in silico cell type deconvolution analysis and built correlation networks to elucidate its role on a transcriptional level. Using our transcriptomic results, we set-up functional experimental assays to understand the effects of OTOF on viral infections in SARS-CoV-2, Human immunodeficiency virus 1 (HIV-1) and Yellow Fever Virus (YFV). Our assays focused on the effects of ectopic OTOF expression on viral binding, entry, replication and exit. In addition, we also performed interferon stimulation assays in primary cells to recapitulate the OTOF transcriptomic pattern observed in whole blood. We observed increased infectivity in cells overexpressing OTOF when subjected to either YFV or HIV-1 but not with pseudotyped SARS-CoV-2 spike proteins tested. Viral binding or fusion remained unaffected in all our assays. Stimulation of CD4+ T cells and macrophages with IFN-α,IFN-β or IFN-γ revealed marked upregulation of OTOF, suggesting its role as an interferon-stimulated gene (ISG). Overall, this thesis provided new insights into the transcriptomic landscape of host immune response upon SARS-CoV-2 infections, taking into account the variability observed with differing ages. We also uncovered new roles for OTOF as an ISG, potentially identifying new antiviral targets in viral infections.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: SARS-CoV-2, ISG, OTOF, Immunology, COVID-19, HIV-1, YFV, Virology, Type I IFN, Genes, Whole blood, RNAseq, Infection, Blam-Vpr, VSV-G
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 23. Juli 2025
• 1. Bericht­erstatter:in: Kim-Hellmuth, Sarah