Stimulation of NEIL2-mediated Oxidized Base Excision Repair via YB-1 Interaction during Oxidative Stress

Abstract

The recently characterized enzyme NEIL2 (Nei-like-2), one of the four oxidized base-specific DNA glycosylases (OGG1, NTH1, NEIL1, and NEIL2) in mammalian cells, has poor base excision activity from duplex DNA. To test the possibility that one or more proteins modulate its activity in vivo, we performed mass spectrometric analysis of the NEIL2 immunocomplex and identified Y box-binding (YB-1) protein as a stably interacting partner of NEIL2. We show here that YB-1 not only interacts physically with NEIL2, but it also cooperates functionally by stimulating its base excision activity by 7-fold. Moreover, YB-1 interacts with the other NEIL2-associated BER proteins, namely, DNA ligase III␣ and DNA polymerase ␤ and thus could form a large multiprotein complex. YB-1, normally present in the cytoplasm, translocates to the nucleus during UVA-induced oxidative stress, concomitant with its increased association with and activation of NEIL2. NEIL2-initiated base excision activity is significantly reduced in YB-1-depleted cells. YB-1 thus appears to have a novel regulatory role in NEIL2-mediated repair under oxidative stress. Reactive oxygen species (ROS)2 are believed to play an important role in inducing various disease pathologies including cancer, rheumatoid arthritis, cardiovascular disease, and aging (1, 2). ROS are formed endogenously as a byproduct of respiration and oxidative metabolism and exogenously by a variety of environmental agents including ultraviolet (UV) radiation (3-5). Of the total ultraviolet light spectrum (100 -400 nm) present in the natural sunlight, only UVA (320 -400 nm) and a small fraction of UVB (280 -320 nm) reach the surface of the earth, as UVB is mostly absorbed by the atmosphere. Although UVA is the predominant genotoxic radiation in the natural environment, it does not react directly with DNA, but interacts with cellular chromophores like riboflavin, porphyrins, quinones, and reduced nicotinamide cofactors to produce singlet oxygen (6). UVA also has the ability to penetrate deeper into the skin to proliferating basal layers, and causes DNA damage leading to genomic instability.Although cellular antioxidant defenses (e.g. catalase, peroxidase, and superoxide dismutase) effectively combat the effects of ROS, but oxidative DNA damage still occurs. Most of the DNA lesions, except double strand breaks, are repaired via the DNA base excision repair (BER) pathway, initiated with the excision of damaged base by a specific DNA glycosylase (7,8). Four oxidized base-specific DNA glycosylases have been identified and characterized so far in mammalian cells. 8-Oxoguanine-DNA glycosylase (OGG1) and endonuclease III homolog 1 (NTH1) were characterized previously and preferentially excise oxidized purines and pyrimidines, respectively (9, 10), and were thought to be the two major oxidized base-specific DNA glycosylases in mammalian cells. However, a lack of phenotype or significant cancer propensity of OGG1-and NTH1-null mice suggested the contribution of other DNA glycosylases in the repair of oxidized ...

Early steps in the DNA base excision/single-strand interruption repair pathway in mammalian cells

“…For example, we have shown ROS-mediated activation of NEIL1 and APE1 [183,184]. We have shown that ROS enhances NEIL2 complex formation [161]. ROS also enhances modification of OGG1 [180] and possibly other BER proteins.…”

“…Formation of the multiprotein complexes for oxidative damage repair is enhanced by ROS [51,161]. It was previously shown that NEILs carry out SN-BER mediated by PNK, Pol β, Lig IIIα and XRCC1 by stably interacting with Pol β, Lig IIIα and XRCC1 [51,115].…”

“…NEIL2 interacts with YB-1, a Y-box binding protein, and it was suggested that YB-1 may be required for the fine-tuning of repair [161]. NTH1 was also shown to interact with and be stimulated by YB-1 [175].…”

Enhancement of NEIL1 Protein-initiated Oxidized DNA Base Excision Repair by Heterogeneous Nuclear Ribonucleoprotein U (hnRNP-U) via Direct Interaction

“…Contrary to the common perception that the repair of oxidatively damaged bases in DNA via the BER pathway involves only a few conserved repair proteins, a more complex picture of BER is now emerging in that BER comprises several alternative subpathways that utilize distinct sets of repair enzymes and other proteins (16,19,21,35,46). We have been exploring the role of noncanonical proteins in oxidized base repair and have characterized the proteins present in immunoprecipitates of early BER enzymes including DGs.…”

Emerging Roles of the Nucleolus in Regulating the DNA Damage Response: The Noncanonical DNA Repair Enzyme APE1/Ref-1 as a Paradigmatical Example

“…Actually, several noncanonical factors have been demonstrated to participate BER, even though their in vivo functions are yet to be fully unraveled. The list of non-BER proteins includes for instance: YB-1 [which has been shown to interact with the endonuclease VIII-like 2 (NEIL2) glycosylase (35), the endonuclease III-like glycosylase (NTH1) and APE1 (29)], NEIL2 [which was also found to interact with the RNA-binding protein hnRNP-U (6)], HMGB1 [which has been implicated in single-strand break (SSB) repair involving Polb (90)] and the tumor suppressor p53, which was also shown to play a role in DNA damage repair through direct binding to APE1 and Polb (168). The list of these non-BER proteins is still growing, supporting the notion that BER in vivo is far more complex than the simple model that we can reconstitute in vitro.…”