Deciphering the functions of protein VII during adenovirus infection.

Abstract

Adenoviruses are double-stranded DNA viruses that cause various common illnesses. Although usually considered self-limiting infections, adenoviruses pose significant risks for immunocompromised individuals. Thus, a comprehensive understanding of adenovirus pathogenesis is crucial for advancing strategies aimed at mitigating adenovirus-related diseases. As adenoviruses are nuclear replicating viruses, they necessarily interact with host chromatin. One way in which adenovirus controls host chromatin function is through the histone-like protein, protein VII. Protein VII is an essential viral protein that packages with and condenses the viral genome within the core of the virion. Much like histone proteins, protein VII contains several post-translational modifications (PTMs) that impact its localization within the nucleus. However, the specific impacts of these PTMs on protein VII function during infection remain unexplored. During late infection when protein VII is abundantly expressed, it localizes to and distorts host chromatin and interacts with several host chromatin-associated proteins, including high mobility group box one (HMGB1). In the nucleus HMGB1 binds to and bends DNA to reorganize chromatin. During times of stress or infection, HMGB1 is released from the cell as an alarmin, where it stimulates an inflammatory response. Protein VII retains HMGB1 on chromatin, yet the mechanism of this interaction and its implications during infection are poorly understood. In human adenoviruses, protein VII (hVII) exhibits high conservation, whereas in different vertebrate adenoviruses, such as murine adenovirus-1 (MAdV-1), protein VII (mVII) shows significant divergence with only 39% similarity to hVII. Despite mVII's known role in viral genome packaging within virions, its specific functions remain largely unexplored. MAdVs serve as a valuable in vivo model for investigating adenovirus pathogenesis, thus understanding how mVII contributes to MAdV infection can elucidate how adenoviruses cause disease.In this thesis, I addressed these gaps in our understanding of protein VII function. I investigated how protein VII interacts with HMGB1 and found that protein VII interacts with HMGB1's A-box and anchors it to chromatin using bacterial two-hybrid and human cell culture assays. Furthermore, I showed protein VII suppresses interferon β induction and exploits HMGB1 for this purpose. To determine how PTMs on protein VII impact adenovirus infection, I created mutant viruses that abrogated or mimicked PTMs on protein VII. I determined that acetylation on protein VII may impact expression of early viral genes, but the PTMs did not appear to impact later stages of infection or localization of protein VII. However, I also discovered that protein VII is likely modified at alternative residues, which may compensate for the mutations we introduced. Finally, I used a chimeric HAdV-5 with a protein VII from MAdV-1, to show that mVII does not directly interact with HMGB1, but HMGB1 is still retained on chromatin during infection. I also discovered that in this chimeric virus, mVII was expressed at lower levels than hVII during infection, resulting in a reduction in viral titers. Additionally, mVII frequently localized to the nucleolus, suggesting a potential divergence in function from hVII. These investigations significantly contributed to our knowledge of the interplay between protein VII and the host nuclear environment and its evolutionary adaptations across diverse adenoviral species.