Toledo Group – University of Copenhagen

Toledo Group

Molecular Aging Program
Center for Chromosome Stability

Research interests

In our recently established lab we are interested in the molecular mechanisms that allow mammalian cells to fully and faithfully replicate their DNA content. In every division cells across species face a plethora of challenges in order to successfully duplicate their genome. Aberrant structures or modifications on the DNA molecule can act as roadblocks that stall and even dismantle replication forks. A lot of focus has been set in the past years on understanding how, in response to these obstacles, cells elicit the so called replication stress responses. Various molecular pathways allow cells not only to restore the damage but to temporarily adapt the replication machinery so that DNA synthesis can be resumed.  

In our lab we focus on understanding how cells have solved a more fundamental level of challenges that are inherent to the architecture of the DNA replication machinery. Specially for higher eukaryotes, DNA replication requires a great level of regulation both locally (at the level of the replisome) and globally (throughout the nucleus) that we still poorly understand. At the replisome multiple proteins act together and cooperate to carry out different functions necessary for DNA synthesis in perfect synchrony. Among these, we want to understand how DNA unwinding is coupled with DNA synthesis, which implies a very different task at the leading and at the lagging strand. Nucleus wide, DNA replication has to be tightly regulated to proceed at multiple sites simultaneously with perfect temporal and spatial coordination. How this is achieved in a three dimensional space so that replication proceeds orderly following the so called replication program is still an outstanding question in the field. When any of these multiple levels of regulation is disrupted, cells can suffer endogenous replication stress, which can increase the rate of genomic aberrations favouring malignant transformation but also lead to irrecoverable DNA damage and cell death. Thus understanding these processes and how cells deal with the different sources of exogenous replication stress is highly relevant for human disease. Importantly, the fundamental challenges that cells encounter during DNA replication can be exploited to develop new strategies to treat diseases like cancer, which is the ultimate goal of our lab. For that, we also carry out small molecule screenings to boost the discovery of novel therapeutic targets that could be used in the treatment of proliferative disorders.

We are a cell biology lab with a taste for modern technologies. We use state of the art methodologies such as high content microscopy to analyse cell biology with quantitative detail in single cell resolution. Our research is performed in transformed and primary human cell lines, where we apply classical and modern genetic manipulation approaches (siRNA and CRISPR), intervention with small molecules, and quantitative mass spectrometry to dissect novel regulatory mechanisms and the key players involved. 

Current project areas

  • Global control of DNA replication and origin firing in mammalian cells
  • Regulators of replication fork stability, fork restart and nascent strand degradation
  • Leading and lagging strand uncoupling as a source of genomic instability
  • Regulation of the endogenous S-phase checkpoint

Selected publications

  • Toledo L, Neelsen KJ, Lukas J. Replication Catastrophe: When a Checkpoint Fails because of Exhaustion. Mol Cell. 2017 Jun 15;66(6):735-749

  • Toledo LI, Altmeyer M, Rask MB, Lukas C, Larsen DH, Povlsen LK, Bekker-Jensen S, Mailand N, Bartek J, and Lukas J. ATR prohibits replication catastrophe by preventing global exhaustion of RPA. Cell, 2013, Nov, 155 (5):1088-1103.

  • Altmeyer M, Toledo LI, Gudjonsson T, Grøfte M, Rask MB, Lukas C, Akimov V, Blagoev B, Bartek J, Lukas J. The Chromatin Scaffold Protein SAFB1 Renders Chromatin Permissive for DNA Damage Signaling. Mol Cell. 2013, Oct 24; 52(2):206-20

  • Toledo LI, Murga M, Zur R, Soria R, Rodriguez A, Martinez S, Oyarzabal J, Pastor J, Bischoff JR, Fernandez- Capetillo O. A cell-based screen identifies ATR inhibitors with synthetic lethal properties for cancer-associated mutations. Nat Struct Mol Biol. 2011 Jun;18(6):721-7.