Systems biology analysis of herpes simplex virus 1 induced host shut-off

• Applicant: Professor Dr. Lars Dölken
• Subject Area: Virology
• Term: from 2015 to 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 272269602

Herpes simplex virus 1 (HSV-1) is the causative agent of the common cold sores but also responsible for severe, life-threatening diseases including encephalitis, pneumonia, hepatitis and generalised skin infections. HSV-1 is a paradigm for virus-induced host shut-off exerted at RNA level. This is thought to be mediated by two viral proteins, namely the virus host shut-off protein (vhs) and ICP27. We employed two cutting-edge methodologies, i.e. 4sU-tagging of newly transcribed RNA and ribosome profiling, to study global changes in transcription and RNA processing as well as their impact on translation throughout the full course of infection. Unexpectedly, we found HSV-1 infection to trigger widespread disruption of transcription termination of cellular but not viral genes. This results in extensive transcription activity bypassing poly(A) sites, commonly extending for tens-of-thousands of nucleotides and into downstream genes. Read-in transcription explained the surprising observation that hundreds of cellular genes were transcriptionally induced but not translated. In contrast to previous reports, we found that HSV-1 does not install a general inhibition of splicing but rather only inhibits post-transcriptional but not co-transcriptional splicing. A manuscript on these fascinating findings is currently in revision with Nature. Work is ongoing to identify the responsible viral gene product. We now seek to study the role of vhs, ICP27 and disruption of transcription termination in HSV-1 induced host-shut-off. This will include the following three work packages:i) We will study co- and post-transcriptional RNA processing and nuclear export by analysing subcellular RNA fractions (total cytoplasmic, nuclear and chromatin-associated RNA).ii) We will detail the role of chromatin status, individual nucleosomes, alterations in DNA-binding proteins and the massive transcription of intergenic regions resulting from disrupted transcription termination in the modulation of cellular and viral gene expression using ATAC-seq (Assay for Transposase-Accessible Chromatin using Sequencing).iii) We will define the interaction of the two key viral proteins mediating HSV-1 host shut-off, namely ICP27 and vhs, with both cellular and viral RNAs at nucleotide resolution using PAR-CLIP (Photo-Activated Ribonucleotide-enhanced Cross-Linking ImmunoPrecipitation). Results obtained from this work will detail a fascinating new host-shut-off mechanism employed by an important human pathogen and establish HSV-1 as an interesting new model to study the molecular mechanisms coupling RNA synthesis, processing, export and translation in mammalian cells.

• DFG Programme: Research Grants