Development of novel nano-agents to target DNA replication forks and modulate cell cycle and activity

New agents that bind to DNA Y-shaped junctions will be studied to further understand the basic cell and molecular biology of these agents in more detail specifically: (i) how they enter the cell and where (and how quickly) they localise, (ii) how they interact with key biomolecules in the cell such as genomic DNA and the downstream cellular effects of those interactions and (iii) the resultant properties in in vitro cellular systems.

Newly modified junction binders (e.g. His-tagged, fluorescently labeled) will be used to image and study cellular interactions and develop novel imaging tools to directly study biologically important DNA transactions (e.g. DNA replication forks) inside of cells by measuring incorporation of fluorescent base analogues and also using isolated DNA under laminar sheer flow. This technique will allow us to analyze the impact of the agents on rates of replication fork progression, frequency of fork stalling or collapse, and also will provide insights into the cellular response to exposure (compensatory origin firing, effect on nearby replication structures). In parallel, we will develop binders that are amenable to cross-linking by 'click' chemistry to identify proteins bound to the DNA within the vicinity of the agent.