"DNA damage (DD) is the cause of several diseases, including cancer, and it is also linked to the organ decline that arises in ageing. Yet, the contribution of different sources of DD to these processes is not understood. Sources of DD such as chromosome breaks, eroded telomeres or oxidative stress are been heavily investigated. For establishing my group, I decided to focus on a source of DD that arises every time a cell replicates its DNA, and which is known as replication stress (RS). In short, RS stands for the excessive accumulation of single-stranded DNA at replication forks that, due to its recombinogenic nature, can initiate genomic rearrangements. Consistently, RS is now known to be a key source of genomic instability in human tumors. In mammalian cells, a signalling cascade initiated by ATR and Chk1 kinases suppresses RS. Unfortunately, the essential nature of these kinases significantly limited the study of the RS-response in mammals. In the initial years of our lab we have developed several tools that facilitate the study of RS in mammals. These include a cellular system where ATR can be activated at will, potent and selective ATR inhibitors, and mice with reduced or increased levels of ATR and Chk1 kinases. These tools have allowed us to start exploring how RS impacts on cancer and ageing, as well as to investigate the potential of targeting ATR for cancer therapy. Yet, the field of RS is still poorly developed, and many basic questions are still in the need of answers.This application outlines a plan for our research in the next five years, and explains how I propose to investigate RS at molecular, cellular and animal levels. Whereas I plan to capitalize on the tools (published and unpublished) that we have generated within the last few years, I also propose several innovative strategies for the study of the RS-response in mammals. This grant would allow us to consolidate our still young group as a solid laboratory for the study of RS in mammals."