Endonuclease VIII-like 3 (Neil3) DNA glycosylase promotes neurogenesis induced by hypoxia-ischemia

Abstract

Neural stem/progenitor cell proliferation and differentiation are required to replace damaged neurons and regain brain function after hypoxic-ischemic events. DNA base lesions accumulating during hypoxic-ischemic stress are removed by DNA glycosylases in the base-excision repair pathway to prevent cytotoxicity and mutagenesis. Expression of the DNA glycosylase endonuclease VIII-like 3 (Neil3) is confined to regenerative subregions in the embryonic and perinatal brains. Here we show profound neuropathology in Neil3-knockout mice characterized by a reduced number of microglia and loss of proliferating neuronal progenitors in the striatum after hypoxia-ischemia. In vitro expansion of Neil3-deficient neural stem/progenitor cells revealed an inability to augment neurogenesis and a reduced capacity to repair for oxidative base lesions in single-stranded DNA. We propose that Neil3 exercises a highly specialized function through accurate molecular repair of DNA in rapidly proliferating cells.DNA damage | formamidopyrimidine-DNA glycosylase/endonuclease VIII | hydantoins | neural stem cells | neuronal progenitor cells T he base-excision repair pathway (BER) maintains genomic integrity by removing base lesions caused by oxidation, alkylation, and deamination. DNA base lesions frequently are cytotoxic or mutagenic if not removed. BER is initiated by DNA glycosylases that recognize modified bases and catalyze cleavage of the N-glycosidic bond, creating an apurinic or apyrimidinic (AP) site. The exposed DNA backbone is cleaved by the AP lyase activity of bifunctional DNA glycosylases or by an AP endonuclease. Repair synthesis is completed by gap filling and ligation (1, 2).Endonuclease VIII-like 3 (NEIL3) and endonuclease VIII-like 1 (NEIL1) are mammalian oxidized base-specific DNA glycosylases (3, 4). The function of NEIL3 has remained enigmatic, but recently the mouse ortholog was shown to remove a broad spectrum of oxidative base lesions on single-stranded DNA substrates with preference for spiroiminodihydantoin (Sp) and guanidinohydantoin (Gh), which are further oxidation products of one of the most common base lesions, 8-oxo-7,8-dihydroguanine (8ohG) (5). These findings suggest that NEIL3 serves as a DNA glycosylase to prevent accumulation of cytotoxic and mutagenic DNA lesions in mammalian cells, although the activity of NEIL1 far exceeds that of NEIL3 on most substrates.In the late postnatal and adult brain, newborn neurons arise from neural stem/progenitor cells (NSPCs) in both the subgranular zone (SGZ) of the hippocampal dentate gyrus and in the subventricular zone (SVZ) (6). We previously reported a discrete expression pattern of Neil3 in the rodent SGZ and SVZ, confined to the embryonic and perinatal stages (7,8). These observations indicate a role for Neil3 in proliferating cells in the brain. However, naïve Neil3-knockout mice generated by us and others (4) appear phenotypically normal. After perinatal hypoxic-ischemic (HI) and adult ischemic stroke, proliferation of SVZ NSPCs is enhanced, and differentiating p...

Oxidative DNA damage in disease—Insights gained from base excision repair glycosylase‐deficient mouse models

“…Hypoxia during ischemia has also been shown to induce neuronal pathology in Neil3 2/2 mice, that display reduced microglial content and a significant loss of neuronal progenitors specifically in the striatum during hypoxic injury [Sejersted et al, 2011]. Furthermore, neural stem/ progenitor cells from aged Neil3 2/2 mice were reported to have reduced proliferative capacity and reduced DNA repair activity, corresponding with learning and memory deficits observed in aged Neil3 2/2 mice [Regnell et al, 2012].…”

Transcriptome profiling of the newborn mouse brain after hypoxia–reoxygenation: hyperoxic reoxygenation induces inflammatory and energy failure responsive genes

“…Since we aimed for a milder oxidative insult, proliferation of microglia increased in a more modest manner as compared with HI in the immature brain (28). Astrocytes are affected by microglia activation after HI, contributing to excessive release of inflammatory mediators which can further intensify the injury (5).…”

“…To determine apoptosis and/or necrosis, a pathologist blinded to the experimental groups assessed hematoxylin and eosin-stained brains in the regions of interest: hippocampus, cortex, and striatum. Immunostaining was performed as previously described (28). The primary antibodies and dilutions used were mouse anti-GFAP 1:400 (G3893; Sigma, St.Louis, MO) and rabbit anti-Iba1 1:2000 (019-19741, Wako, Richmond, VA).…”

The endonuclease VIII-like proteins: new targets in the treatment of ischemic stroke?

“…NEIL3 expression has also been detected in regions rich in neurogenesis in the embryonic brain [28] and in two recent studies NEIL3-null models showed a decreased differentiation potential of neural stem cells. [29,30] NEIL3-null mice showed learning and memory deficits and reduced anxiety-like behavior, and synaptic irregularities in hippocampal neurons. [31] All together, these results suggest that NEIL3 may have a specific role in neurogenesis in the central nervous system.…”

“…[36] In order to understand the relationship between NEIL3 and hypoxia-ischemia, Sejersted et al [29] carried out an experiment on NEIL3 gene knockout (NEIL3-null) mice in vitro and in vivo. Interestingly, after hypoxia-ischemia, there is no increase of cellular damage or death in vivo in NEIL3-null mice at an early stage, but a significant deficient in reconstituted neuronal tissue after 42 d. NEIL3-null neurospheres exhibited poor growth and skewed differentiation that could explain the poor outcome of NEIL3-null mice after hypoxic ischemia.…”