CBP and p300 acetylate PCNA to link its degradation with nucleotide excision repair synthesis

Abstract

The proliferating cell nuclear antigen (PCNA) protein serves as a molecular platform recruiting and coordinating the activity of factors involved in multiple deoxyribonucleic acid (DNA) transactions. To avoid dangerous genome instability, it is necessary to prevent excessive retention of PCNA on chromatin. Although PCNA functions during DNA replication appear to be regulated by different post-translational modifications, the mechanism regulating PCNA removal and degradation after nucleotide excision repair (NER) is unknown. Here we report that CREB-binding protein (CBP), and less efficiently p300, acetylated PCNA at lysine (Lys) residues Lys13,14,77 and 80, to promote removal of chromatin-bound PCNA and its degradation during NER. Mutation of these residues resulted in impaired DNA replication and repair, enhanced the sensitivity to ultraviolet radiation, and prevented proteolytic degradation of PCNA after DNA damage. Depletion of both CBP and p300, or failure to load PCNA on DNA in NER deficient cells, prevented PCNA acetylation and degradation, while proteasome inhibition resulted in accumulation of acetylated PCNA. These results define a CBP and p300-dependent mechanism for PCNA acetylation after DNA damage, linking DNA repair synthesis with removal of chromatin-bound PCNA and its degradation, to ensure genome stability.

P300 inhibition enhances gemcitabine-induced apoptosis of pancreatic cancer

“…Because p300 in the nucleus has been shown to exist as either a soluble form or a chromatin-bound form [17], we separated the nuclear protein into a soluble fraction and a chromatin-bound fraction. Although the chromatin-bound form of p300 represented a much smaller proportion of the entire nuclear p300 pool, the chromatin-bound p300 is likely to be important in the DNA damage response (Figure 2A).…”

“…Consistent with our observations in the present study, p300 was shown to be recruited at the sites of DNA break induced by UV or IR in human cells and facilitate DNA repair. Although p300 cannot promote DNA repair by itself, p300 has been considered to serve as a cofactor for multiple mechanisms of DNA repair synthesis [6, 17, 23, 24]. For instance, proliferating cell nuclear antigen (PCNA) is known to accumulate at the radiation-induced DNA damage sites and promote subsequent DNA repair.…”

“…Since the spectrum of target residues for p300 in histone proteins is wide and overlapped by the target residues by other proteins with HAT activity, it is not practically possible to identify the exact histones residue responsible for p300-mediated protection from gemcitabine-induced DNA damage. While the conformational change of the chromatin with acetylation of histones by p300 and the subsequent the recruitment of various DNA binding cofactors have been implicated as the primary mechanism of p300-mediated DNA repair [6, 17, 23, 24], Ogiwara and colleagues described other possible mechanisms by which p300 may also influence DNA repair. They reported p300-mediated transcriptional activation of DNA repair genes including BRCA1 and RAD51, upon IR induced DNA damage [23].…”

Defective DNA repair and increased chromatin binding of DNA repair factors in Down syndrome fibroblasts

“…In agreement with these conclusions, it has been recently reported that excessive retention of DNA replication and repair factors on chromatin is dangerous for genome stability [46,47]. In particular, mutant forms of PCNA protein that could not be disassembled from chromatin, showed a reduced DNA repair efficiency [48]. Future investigations are needed to elucidate the link of this phenomenon with DS in order to fully clarify the molecular basis of genetic instability in this disease.…”

The interaction between ALKBH2 DNA repair enzyme and PCNA is direct, mediated by the hydrophobic pocket of PCNA and perturbed in naturally-occurring ALKBH2 variants

“…We also note a slower migrating PCNA band that copurifies with the ALKBH2 Q10K variant. While the exact nature of the higher molecular weight PCNA band is currently unknown, the additional PCNA species could represent an acetylated form of PCNA based upon its migration [24]. By using a restriction enzyme-mediated AlkB DNA repair assay, the ALKBH2 variants display comparable enzymatic activity to WT ALKBH2 (Figure 3C and D).…”