Epitranscriptomic regulation of viral mRNAs during herpes simplex virus type 1 infection

• Applicant: Dr. Denise Ohnezeit
• Subject Area: Virology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 554758329

The proposed research project investigates how viral mRNAs are regulated epitranscriptomically by N6-methyladenosine (m6A), an abundant and ubiquitous chemical RNA modification. Because recent advances in high-resolution sequencing approaches have reignited the field of epitranscriptomics, the project will be at the forefront of this field by applying highly innovative technologies to study m6A on a genome-wide, but transcript- and isoform-level, with quantitative, high-precision and base-level resolution. My goal is to rigorously evaluate and improve existing m6A detection technologies and to create novel m6A-manipulation strategies to investigate for the first time, the biological consequences of m6A during infection of relevant cell types with herpes simplex virus type 1 (HSV-1). The first work package (WP) comprises an in-depth comparison of different orthogonal m6A mapping techniques and their performance on HSV-1 transcripts. WP2 will focus on the biological relevance and the dynamics of m6A during HSV-1 infection. A major goal is to compare the m6A methylome maps of HSV-1 transcripts from normal human dermal fibroblasts (NHDFs) over the course of a lytic infection and reveal if m6A deposition on newly made viral transcripts differs over time. This is relevant because the virus expresses a regulatory protein ICP27 that reduces the availability of the methyltransferase complex during the later stage of infection. Furthermore, by creating m6A methylome maps in a human neuronal cell line, investigating both lytic infection and reactivation from latency, I expect to see decisive changes in the occupancy of m6A sites, consistent with preliminary data indicating a dependency on m6A for viral genome replication in these cells. In WP3, I will develop a highly specific CRISPR-Cas-based tool to target m6A sites for directed methylation or demethylation. This will enable me to target m6A sites of interest on selected transcripts in live infected cells. Because of its innovation and high specificity, this strategy will be applicable to any viral or host transcript of interest and has the potential to reveal as yet unidentified consequences of m6A for the fate of individual transcripts. This exciting project brings together highly innovative and orthogonal methods and will help to close major gaps in our knowledge of epitranscriptomic regulation of highly overlapping and dense viral genomes, using HSV-1 infection as an impactful model. Further, the evaluation and improvement of novel technologies will be important to the wider scientific community, either in host-pathogen interactions or human diseases, including cancer, and will serve as a launch point for my future scientific career after the two-year stay at New York University.

• DFG Programme: WBP Fellowship
• International Connection: USA
• Host: Professor Ian Mohr, Ph.D.