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Title: Integrative analysis of transcriptional activity and genome architecture changes upon viral infections
Author: Michalski, Marco Alexander
ISNI:       0000 0004 7228 3935
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2018
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To study the interplay between spatial nuclear architecture and transcriptional activity during viral infections, I employed a genome-wide chromosome conformation capture approach (Hi-C) on infected murine and human cells and further enriched those libraries for genomic loci of interest and the viral genomes with biotinylated RNA baits. In parallel, I profiled newly transcribed RNA throughout the entire kinetic of murine cytomegalovirus (mCMV) infection in mice. Host genome rearrangement is a well-known phenomenon of mCMV infection but the underlying mechanisms are largely unknown. Furthermore, HPV infection can lead to cervical cancers in humans, with genomic instability and re-arrangements, leading to dysregulation of gene expression. Thus studying changes in genome architecture at early stages of HPV induced carcinogenesis can further our understanding on how certain integration events can provide a growth advantage. In this study, I identified clusters of genes characterized by distinct kinetic profiles upon CMV infection in the mouse, which were associated with distinct functional terms. ATAC-Seq uncovered proximal promoter regions (PPR) that showed an over-representation of specific transcription factor binding sites in each of the clusters. These correlated well with the annotated functions of the associated clusters. Further, I found that lytic mCMV infection is accompanied by local and global changes of chromosomal interactions in the host cell genome. Notably, chromatin properties, such as gene density, GC content and the association with the nuclear lamina, predict the structural dynamics upon infection and correlate well with transcriptional activity and changes thereof. High-resolution interaction profiles for TSSs of highly induced or repressed genes, suggest that in general, enhancer-promoter interactions already form in untreated cells; and these pre- existing DNA-structures are not significantly altered but function through transient activation or repression of enhancers. Finally, the viral genome showed a distinct pattern of open and closed chromatin late in infection. We found that the 7.2 kb viral intron displays the most open chromatin, and is highly enriched for chromosomal contacts with the host genome. Hi-C and capture Hi-C revealed that both short- (~50 kb) and long-range (~1 Mb) interactions occur during the early stages of HPV induced carcinogenesis between the host and the integrated HPV16 genomes. Integration and direct interactions between the viral genome and the host DNA were shown to be associated with changes in host gene expression. In addition, insertion of the virus can disrupt normal host architecture. In summary, this project pioneers the study of changes in nuclear architecture upon viral infection in man and mice. I uncover numerous structural features and changes of both the viral genomes and the infected host cellular genomes, and I demonstrate that these changes correlate with transcriptional activity.
Supervisor: Fraser, Peter Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: Hi-C ; mCMV ; HPV-16