DNA Damage Response to double-stranded DNA break -- Homologous Recombination v 4.0- Full HD
The DNA damage response (DDR) of the cell includes: (i) sensing, (ii) signalling and (iii) repair of such damage. Double strand breaks (DSBs) are the most toxical DNA damage for the cell. They can be induced by ionizing radiation, laser beam, bleomycin, Topoisomerase II enzyme, endonucleases or also can be produced during the repair of single-stranded breaks (SSB) DNA. It has been reported that approximately nine DSB per cell and day are produced in physiological conditions1. The Homologus Recombination (HR) pathway is in charge of DSBs repair, in a error-free fashion, during S or G2 phases of the cell cycle, by using sister chromatid as template3. A proficient sensing and signalling of DSBs is very important for the maintenance of the genome and chromosomal stability.
Recent research works, stressed out the key role of post-traslational modification of DDR proteins such as: phosphorilation, acetylation, methylation, ubiquitination and sumoylation in regulating the DNA damage signalling and response. The HR DDR signalling is believed to act in the following order: first, the DSB lesion is recognized by MRN complex (MRE11-RAD50-NBS1), that recruits the ATM (mutated in Ataxia Telangiectasia) kinase into the damage site. ATM phosphorilates the serine 139 of the γH2AX at the damage site and also in large number of nucleosomes around the DSB. ATM also phosphorilates MDC1 (mediator of DNA damage checkpoint protein 1). At this point the γH2AX-ATM-MDC1 connection generates a possitive feedback loop, that contributes to amplify DSB signal along the whole nucleous. Following this, RNF8 (an ubiquitin ligase), modifies γH2AX. Then RNF168, an E3 ubiquitin ligase, detects the RNF8 signal in histones and amplify it by creating poly-ubiquitin chains. At this time, the RAP80-Abraxas complex is recruited into the damage site, followed by the BRCA1-BRCC36 complex. BARD1 it is believed to win the competition against Ctip protein for coupling with BRCA1-BRCC36 complex. An important role is attributed to RAP80 ubiquitination by RNF8, and BRCA1 SUMOylation by PIAS1-UBC9 complex. At this time, Ctip couples to MRN complex, displace most of the DDR proteins at DSB site, and catalyzes the DSBs ends resection by using its exonuclease activity. Immediately RPA binds to ssDNA. Then BRCA2 displaces RPA and enhance the RAD51 binding to the ssDNA filaments. This is the main signal for recruitment of recombination machinery to complete the DSB repair.