Untangling the genetic wiring behind the activation of immune and inflammatory responses

• Applicant: Sara Becker, Ph.D.
• Subject Area: Virology ; Immunology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 546121141

Interferons (IFNs) are a class of cytokines that were originally described as antiviral proteins. However, with time, it became evident that they are involved in multiple key processes within the innate immune response. IFNs trigger the expression of IFN-stimulated genes (ISGs), which induce an antiviral state in cells neighboring infected cells. The expression dynamics of these ISGs are managed through a complex network of different enzymes and transcription factors. These networks are only partially understood, especially when it comes to understanding which transcription factor is responsible for the expression of a certain gene at distinct timepoints. To answer this, we will set up a systematic analysis of transcriptional processes of human epithelial cells after IFN treatment. Additionally, we compare these to the transcriptome induced directly by viral infections. This will allow the identification of common pathways of innate antiviral responses, but also the perturbations viruses induce through immune evasion mechanisms. As a first step, we will analyze the early transcriptome of IFNβ treated human epithelial cells through RNA sequencing (RNAseq) at different timepoints. With this, we will construct gene regulatory networks (GRNs) using single-cell regulatory network inference and clustering (SCENIC). These networks will be the prerequisite to cluster ISGs depending on their expression kinetics. We will then characterize these different clusters by analyzing their promoter and enhancer regions to find common patterns of transcription factor binding. As there are already many RNAseq datasets available on public databases, suitable datasets will be used to compare and complement our analyses. To refine distinct pathways in our GRNs, a pooled single-cell CRISPR screen will be performed. Additionally, distinct genes of interest will be knocked out using CRISPR technology to further elucidate their role in the antiviral response. In a parallel approach, we plan to apply a similar workflow to map GRNs during viral infection. Here, we will use a panel of different viruses to infect susceptible cells and analyze their transcriptome. This analysis will reveal the shared antiviral responses across diverse virus families and identify immune reactions frequently circumvented by viruses. With this proposal, we aim to set up a systematic analysis of transcriptional processes in the IFN-response. We will clarify the expression kinetics of ISGs and further elucidate their role in antiviral responses. In the future, this established workflow can be easily applied to different types of IFN and cell types. Ultimately, these analyses will improve our understanding of the initiation of innate immune responses, and hopefully lead to novel treatment options for not only viral infections, but also autoimmune disorders and cancer.

• DFG Programme: WBP Fellowship
• International Connection: USA
• Host: Professor Daniel Blanco-Melo, Ph.D.