Regulation of resection by chromatin associated proteins
- Autores: Fernando Mejías Navarro
- Directores de la Tesis: Pablo Huertas Sánchez (dir. tes.)
- Lectura: En la Universidad de Sevilla ( España ) en 2019
- Idioma: español
- Número de páginas: 320
- Tribunal Calificador de la Tesis: Teresa Roldan-Arjona (presid.), Ana Beatriz García Rondón (secret.), Felipe Cortés Ledesma (voc.), Mónica Pradillo Orellana (voc.), Marta Martín Flix (voc.)
- Programa de doctorado: Programa de Doctorado en Biología Molecular, Biomedicina e Investigación Clínica por la Universidad de Sevilla
- Materias:
- Enlaces
- Tesis en acceso abierto en: Idus
- Resumen
The DNA molecule carries the genetic information of every living organism but it is permanently exposed to multiple endogenous and exogenous sources of damage that threaten the genomic integrity of its sequence. Among the insults affecting the DNA, one of the most dangerous lesions are the Double strand breaks (DSBs), because they involve a total interruption of the sequence in both strands and thus favour gross chromosome rearrangements. For this reason, if breaks remain unrepaired or missrepaired, it can lead to several diseases including cancer. To overcome this potential risk, cells have evolved two main alternative mechanisms to repair the DSBs: Non-Homologous End-Joining (NHEJ) and homologous recombination (HR) pathways. The balance between both pathways is a capital issue to ensure the proper restoration of the original DNA sequence, but regulation of this process is not fully understood. Among factors that influence the NHEJ/HR ratio, like cell cycle checkpoints or resection process, chromatin also plays an important role modulating the response to DNA damage. Chromatin is not static but is a dynamic structure that wraps and protects the DNA molecule from degradation but also modulates the accessibility of the repair machinery to the lesion. In this Thesis, we selected several chromatin associated proteins to study them as possible candidates for the regulation of the NHEJ/HR balance in human cells, taking advantage of a fluorescent repair system developed in our laboratory that measures the ratio between both repair pathways, the so-called SSR2.0 system. First, we describe a new role of the RAD51 paralogs in earlier steps of DSB repair than those described before, helping RAD51 recombinase to form a nucleoprotein filament and in the Holliday Junction structure resolution during the HR pathway. Specifically, all five RAD51 paralogs seem to hamper the recruitment of the NHEJ machinery, mainly the DNA-PK complex, to resected DNA to promote the HR pathway in detriment of the NHEJ one. Second, we show that the known chromatin remodeler and transcription factor ALC1 controls the DNA resection process together with the translation factor eIF4A1, independently of its helicase activity or its ability to be recruited to damaged DNA. Strikingly, ALC1/eIF4A1 regulate mRNA stability of CtIP, the key factor for resection promotion and subsequent HR pathway success. This regulation is based on the presence of G-quadruplex (G4) structures in the 5’UTR region of one of the two CtIP mRNA isoforms that encode for the canonical CtIP protein. This kind of structure is specifically recognised by the protein eIF4A1. The solely presence of this G4 confers stability modulation depending on the ALC1/eIF4A1 function. Lastly, we have reported a nuclear direct role of ELP6 protein in DSB repair that is independent of the histone acetyltransferase (HAT) activity of the transcription elongator complex but dependent of the chromatin context. Moreover, maybe this phenotype is shared with the subunits ELP4 and ELP5 that form the hexaremic ring subcomplex. Taking all the data from this Thesis, we set for the first time new roles of chromatin associated factors that affect the regulation of the choice between the two main DSB repair pathways. Further investigations will be required to clarify all the mechanisms involved in these processes and their implications in desease and cancer therapy.
