Main Research Lines:
- Mechanistic analysis of DNA damage signaling upon replication of damaged DNA template in human cells.
- Understanding the process of DNA damage tolerance during replication.
Main Research Lines:
My research career has mainly focused on understanding how cells maintain the stability of their genome, specifically during the process of DNA replication.
In 2007, I was awarded with a competitive 4-year FPI fellowship from the University of Sevilla/El Monte foundation to start my PhD at the Andalusian Molecular Biology and Regenerative Medicine Centre (CABIMER, Sevilla) under the supervision of Dr. Ralf Wellinger. During this time, I carried out a self-dependent investigation on the role of manganese homeostasis on genome instability using yeast as a model organism (García-Rodríguez et al, JBC, 2012 and 2015). Furthermore, in collaboration with Prof. Andrés Aguilera (CABIMER, Sevilla), Ruth Stuckey and I discovered an unexpected role of RNA-DNA hybrids in origin-independent replication initiation within the ribosomal DNA locus (Stuckey et al, PNAS, 2015). I also carried out short-term research visits in the prestigious lab of Prof. Rodney Rothstein (Columbia University, New York City, USA) for three months in 2009, performing a high-throughput genetic screen, and in the lab of Prof. Marc Blondel (University of Brest, Brest, France) for two weeks in 2012, developing a yeast-based genetic screen for chemical compounds active against Hailey-Hailey disease.
After finishing my PhD, I joined in 2013 the laboratory of Professor Helle Ulrich, a recognized expert on the field of DNA damage and repair, at the Institute of Molecular Biology (IMB, Mainz, Germany), where I became interested on exploring how yeast cells sense and deal with DNA damage during replication During this postdoc stage, I discovered a novel mechanism of checkpoint activation upon replication stress in yeast cells, which involves resection at ssDNA gaps behind replication forks (EMBO J, 2018). This article merited a comment article on the same journal. I also uncovered a new function of the helicase Pif1 in the template switching pathway of DNA damage bypass, signing the article as corresponding author (NAR, 2018). Furthermore, I was involved in other projects or collaborations expected to render high impact publications (manuscripts in prep.), and I also produced a review article and a book chapter on the DNA replication and replication stress topics (Front Genet, 2016; Methods Enzymol, 2019).
In 2018, I was awarded with a Marie Curie Individual Fellowship to continue my work at the lab of Dr. Pablo Huertas at CABIMER. As Principal Investigator of the project and taking into consideration the knowledge generated previously in yeast, I am now analyzing the mechanism of checkpoint activation and DNA damage tolerance during replication of damaged template on human cell lines by applying a multidisciplinary approach, which includes the analysis of stretched single DNA molecules or the generation of true knockout cell lines by CRISPR/Cas9 technology. The results obtained during the course of this project are expected to enhance our understanding of how human cells perceive and respond to DNA damage during replication, a process that is highly relevant to cancer development.
As Principal Investigator:
The genetic information encoded by DNA is under constant attack from both endogenous and exogenous sources of damage. To ensure genome stability and prevent disease, cells use global signaling networks to sense and repair DNA damage. One particularly serious problem is when the replication machinery encounters lesions remaining in the template DNA. In this scenario, cells employ damage bypass mechanisms to complete genome replication and prevent fork breakage. Importantly, these pathways are not restricted to the site of stalling but can also function behind the fork at single-stranded DNA (ssDNA) gaps originated by the re-priming of DNA synthesis downstream of lesions. While it is very well known that ssDNA is the molecular signal that triggers the checkpoint response, it is less clear how and where ssDNA actually arises. Generally, it is assumed to accumulate at stalled replication forks, either by an uncoupling between replicative helicase and polymerase movement or between leading and lagging strand synthesis. However, in a recent study in budding yeast, I found that ssDNA gaps left behind replication forks, and extended by processing factors such as the exonuclease EXO1, constitute the predominant signal that leads to checkpoint activation in response to damaged DNA templates during S phase. Whether this mechanism of checkpoint activation is conserved from yeast to humans remains unexplored. Hence, using a unique set of multidisciplinary approaches, this project aims to address the fundamental question of how DNA damage is sensed during replication in human cells. Interestingly, not only ssDNA gap processing seems important for checkpoint signaling but also for the template switching mechanism of damage bypass. Therefore, this project will also study the function of EXO1 and its association with PCNA at postreplicative ssDNA gaps in order to shed light on the poorly understood mechanism of template switching.