Nanopore Direct RNA Sequencing and Proteomics Reveals Virus-induced Changes in Human Cells

Abstract

Influenza virus remains a pervasive global health threat due to its rapid mutation and immune evasion capabilities, resulting in frequent epidemics and sporadic pandemics. This thesis investigates the influence of N6-methyladenosine (m6A) RNA modifications on host-pathogen interactions during influenza infection, emphasizing their role in regulating gene expression and host cellular processes. Leveraging nanopore direct RNA sequencing and proteomic analyses, this study offers a comprehensive view of the epitranscriptomic changes in influenza-infected human cells.The research explores the global distribution of m6A modifications across host and viral RNA, focusing on their effects on RNA stability, splicing, and translation. Through proteomics integration, we assess how these epitranscriptomic modifications influence protein expression and cellular function, elucidating the contribution of m6A modifications and m6A-binding proteins to immune response regulation and viral replication control. The study identifies a dynamic role for influenza-induced m6A modifications, which can both support antiviral signaling and, conversely, facilitate viral replication. This dual functionality reveals an intricate balancing act between viral manipulation of host pathways and host adaptive responses aimed at containment. Key findings underscore that influenza infection triggers specific m6A modifications, enhancing the stability and translation of immune-related transcripts, thereby bolstering antiviral defenses. Additionally, a conserved m6A consensus motif (GGACU) was observed, indicating a stable targeting mechanism for m6A methylation across host and viral RNA, a feature maintained even under infection-induced stress. This conservation suggests that while infection modulates m6A distribution and intensity, it does not disrupt its fundamental targeting, allowing m6A regulation of host defenses to persist. The research highlights the therapeutic potential of targeting m6A modifications, suggesting that precision interventions within this pathway could provide new strategies for antiviral therapies. By mapping the roles of m6A in virus-host interactions, this study contributes to the broader understanding of epitranscriptomic regulation in viral infections, with implications for developing antiviral therapies that harness RNA modification pathways.