HECTD1 promotes base excision repair in nucleosomes through chromatin remodelling.
Nucleic Acids Res
; 48(3): 1301-1313, 2020 02 20.
Article
em En
| MEDLINE
| ID: mdl-31799632
ABSTRACT
Base excision repair (BER) is the major cellular DNA repair pathway that recognises and excises damaged DNA bases to help maintain genome stability. Whilst the major enzymes and mechanisms co-ordinating BER are well known, the process of BER in chromatin where DNA is compacted with histones, remains unclear. Using reconstituted mononucleosomes containing a site-specific synthetic abasic site (tetrahydrofuran, THF), we demonstrate that the DNA damage is less efficiently incised by recombinant AP endonuclease 1 (APE1) when the DNA backbone is facing the histone core (THF-in) compared to that orientated away (THF-out). However, when utilizing HeLa whole cell extracts, the difference in incision of THF-in versus THF-out is less pronounced suggesting the presence of chromatin remodelling factors that stimulate THF accessibility to APE1. We subsequently purified an activity from HeLa cell extracts and identify this as the E3 ubiquitin ligase, HECTD1. We demonstrate that a recombinant truncated form of HECTD1 can stimulate incision of THF-in by APE1 in vitro by histone ubiquitylation, and that siRNA-mediated depletion of HECTD1 leads to deficiencies in DNA damage repair and decreased cell survival following x-ray irradiation, particularly in normal fibroblasts. Thus, we have now identified HECTD1 as an important factor in promoting BER in chromatin.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA
/
Ubiquitina-Proteína Ligases
/
Reparo do DNA
Tipo de estudo:
Prognostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article