Andrés Clemente Blanco

Cell cycle and genome stability

Endogenous metabolic products, such as reactive oxygen species, and exogenous physical and chemical genotoxic stress, constantly assault the genetic material in the cell. It has been estimated that there are about 105 lesions per cell per day in humans. In response to such a high levels of DNA damage, sophisticated mechanisms have evolved to coordinate cell cycle progression with DNA repair. When those systems fail or the rate of DNA damage exceeds the capacity of the cell to repair it, the accumulation of errors can overwhelm the cell and result in early senescence, apoptosis, or cancer.

Recently, a rapid progress has been made in defining the cellular response to DNA damage. After a DNA damage has been produced, a DNA damage response (DDR) signal is triggered in order to couple DNA repair with cell cycle progression. This signal is driven by phosphorylation events, (predominantly serine and threonine residues) that activate both the repair of the broken DNA and the DNA damage checkpoint arrest. While the importance of several kinases in the DDR has been well documented over the last years, the complex role of protein dephosphorylation has only begun to be investigated. Surprisingly, different studies have pointed forward the essential function of several phosphatases in DNA repair, not only by counteracting the DNA damage-induced phosphorylation events to promote reentry in the cell cycle, but also by playing a direct role in the repair process.

Taking in to account the great number of techniques and tools available in Saccharomyces cerevisiae in order to dissect different aspect of DNA repair, it is tantalizing to use this system to unravel new functions of different protein phosphatases in response to DNA damage. One of the best-described models to analyze DNA repair in S. cerevisiae is the homing endonuclease HO system. Since the HO is a site-specific endonuclease, we can explore the repair of a single DNA cut using standard genomic assays, gathering information about how the broken DNA is repaired by the cell (see Figure). By using this methodology and other molecular biology techniques, our goal is to understand how protein phosphatases are involved in the repair of a DNA lesion, their regulation during the activation of the DNA damage response and their targets in the process, in o order to obtain a complete picture of the role of these proteins in the maintenance of genome stability.

Analysis of DNA repair

A) Upon generation of a double strand break (DSB) cells can repair the DNA lesion by using homologous recombination to generate both gene conversion and crossover products. B) The different DNA repair outcomes can be detected by using standard Southern blot assays. C) The Cdc14 phosphatase, together with the DSB itself, are recruited to adjacent areas of the SPBs (Spindle pole bodies) to enhance the repair of the DNA lesion. D) In response to DNA damage, the phosphatase activity of Cdc14 is vital for stabilizing the metaphase spindle and promoting the recruitment of DNA lesions to the SPBs to favour their repair.

Group members
Andrés Clemente-Blanco Principal Investigator
Facundo Nehuén Ramos PhD student
María Teresa Villoria PhD student
Pedro Charria Undergraduate student
Eva Merino García Technician
Recent publications
Villoria MT, Ramos F, Dueñas E, Faull P, Cutillas PR, Clemente-Blanco A (2017).
Stabilization of the methaphase spindle by Cdc14 is required for recombinational DNA repair.
EMBO Journal. 4;36(1):79-101.
Ramos F, Leonard J, Clemente-Blanco, Aragón L (2017).
Cdc14 and chromosome condensation: Evaluation of the recruitment of condensin to genomic regions.
Methods Mol Bio. 1505:229-243.
Bermúdez-López M, Villoria MT, Esteras M, Jarmuz A, Torres-Rosell, Clemente-Blanco A, Aragón L (2016).
Sgs1´s roles in DNA end resection, HJ dissolution, and croosver suppression require a two-step SUMO regulation dependent on Smc6/6.
Genes Dev. 1;30(11):1339-56.
García-Luis J, Clemente-Blanco A, Aragón L, Machín F (2014).
Cdc14 targets the Holliday junction resolvase Yen1 to the nucleus in early anaphase.
Cell Cycle. 13:1392-9.
McAleenan A*, Clemente-Blanco A*, Cordon-Preciado V, Sen N, Esteras M, Jarmuz A, Aragon L (2013)
Post-replicative repair involves separase-dependent removal of the kleisin subunit of cohesin.
Nature. 493:250-254.
Research grants
MINECO: BFU2016-77081-P
MINECO: BFU2013-41216-P
Links of interest
Saccharomyces genome database