Oxidative Stress Sensing and Response in Neural Stem Cell Fate

Abstract

Neural stem/progenitor cells (NSPCs) contribute to the physiological cellular turnover of the adult brain and make up its regenerative potential. It is thus essential to understand how different factors influence their proliferation and differentiation to gain better insight into potential therapeutic targets in neurodegenerative diseases and traumatic brain injuries. Recent evidences indicate the roles of redox stress sensing and coping mechanisms in mediating the balance between NSPC self-renewal and differentiation. Such mechanisms involve direct cysteine modification, signaling and metabolic reprogramming, epigenetic alterations and transcription changes leading to adaptive responses like autophagy. Here, we discuss emerging findings on the involvement of redox sensors and effectors and their mechanisms in influencing changes in cellular redox potential and NSPC fate.

Graphical Abstract

1. Introduction

The persistence of neural stem/progenitor cells (NSPCs) ensures the regenerative potential of the adult brain [1]; [2]. NSPC fate changes depending on pathophysiological variables [3]. Disease conditions can involve defective antioxidant systems and radiation, [4], metabolism [5], and inflammation [4]; [6]. Aging, in particular, influences NSPC fate [7]. Biologically, the cellular milieu of the aging brain exhibits increased signs of impaired bioenergetics and adaptive neuroplasticity, aberrant neuronal activity and calcium homeostasis, the accumulation of oxidatively damaged molecules and organelles, and inflammation [8]. The brain is particularly vulnerable to oxidative stresses due to its high oxygen consumption, polyunsaturated fatty acids enriched in the neuronal membrane, and the presence of excitotoxic amino acids [9].

Reactive oxygen species (ROS) are unstable and reactive derivatives of oxygen such as hydroxyl radicals (•OH), superoxide anion radicals (O2•-), hydrogen peroxide (H₂O₂) and peroxynitrite (ONOO-) in a cell. These are by-products of cellular activities like mitochonrial respiration and fatty acid oxidation which contribute to the oxidative stress [10]. The major sources of ROS are cellular organelles including mitochondria, peroxisomes, endoplasmic reticulum, and lysosomes as well as enzymatic reactions like those of NADPH oxidases (NOXs) and cyclooxygenase [10]. Cellular signal transduction, such as activation of receptor tyrosine kinases, also increases cellular ROS levels [11, 12]. Therefore, NSPCs have different levels of ROS based on cellular contexts like proliferation and differentiation.

The cellular redox state also regulates the balance between the self-renewal and differentiation of NSPCs (Figure 1). Redox signaling has been found to play a significant role in regulating differentiation of NSPCs and clonal expansion in their proliferative niches [13–16]. Under normal physiological conditions, the production of ROS is tightly regulated by the anti-oxidative scavenging system to maintain stemness properties of the NSPCs. Excessive levels of ROS accumulate and lead to oxidative stress conditions when ROS generation outpaces the scavenging system. High level of ROS can induce NSPC death and quiescence [13, 17]. On the other hand, elevated ROS are associated with increased production of their immediate progenitors [18]. Growth signaling-activated proliferative and multipotent NSPCs were shown to maintain a high ROS status for self-renewal and neurogenesis [14]. For example, NOX promotes maintaining self-renewal of NSPCs [14, 19]. Pharmacological or genetic studies showed NOX2 or NOX4 are necessary for NSPCs proliferation by inducing H₂O₂ production and PI3K/AKT pathway activation, suggesting that the basal ROS production is required for proliferation of NSPCs [14, 20, 21]. More recently, quiescent adult hippocampul NSPCs have been characterized to maintain high intracellular ROS levels which causally decrease as cells become activated [13]. Physiological stimuli induced a NOX2-mediated transient surge of ROS for the transition to activation, a finding emphasizing the dynamic coordination of ROS levels and NSPC fate. The reversible balance between NSPC quiescence and activation is important for adult neurogenesis and ROS dynamics surely matter in this fate decision.

Figure 1. Redox potential determines NSPCs fate.

Schema for NSPC responses upon redox potential changes. ER: endoplasmic reticulum, NOX: NADPH oxidase, COX: cyclooxygenase, LOX: lipoxygenase, FA oxidation: fatty acid oxidation, PPP: pentose phosphate pathway, GPx: glutathione peroxidase, Trx: thioredoxin, Prx: peroxiredoxin

A number of genetic and pharmacological strategies were employed to reach the consensus that redox homeostasis and stress steer NSPC fate. Here, we will review current understanding of redox sensors and effectors and their mechanisms of action on NSPC fate change. We will further discuss redox signaling intermediates and redox responsive transcription factors as direct sensors and effctors of redox challenge, and ROS-induced epigenetic alterations, activation of autophagy, and metabolic reprogramming as adaptive responses to cope with redox-stress in NSPCs (Figure 1).

2. Redox Sensors and Effectors in NSPCs

Redox sensing is achieved by the biochemical and molecular alterations on specific sensor molecules. Intracellular ROS exist either as the reactive superoxide anion or its reduced hydrogen peroxide and hydroxyl radicals which can participate in reduction–oxidation-dependent signaling mechanisms [22]. The list of redox sensors is in Table 1. NSPCs respond to redox changes in the form of proximal signaling, intermediate epigenetic alterations and adaptive gene expression leading to reconfiguration of organelles and longer-term changes. We will discuss these changes characterized in NSPCs below.

Table 1.

The list of redox-sensing proteins

Molecule	Family/Class	Modification	Effect of the modification	Reference
AMPK	Kinase	Phosphorylation	Activation	[23]
ATM	Kinase	Phosphorylation	Activation	[24]
ASK1	Kinase	Phosphorylation	Activation	[25]
ERK	Kinase	Phosphorylation	Activation	[26]
IKK	Kinase	Phosphorylation	Activation	[27]
		Cysteine oxidation	Inactivation	[28]
JNK	Kinase	Phosphorylation	Activation	[29]
P38 MAPK	Kinase	Phosphorylation	Activation	[30]
mTORC1	Kinase	Cysteine oxidation	Activation	[31]
PI3K/AKT	Kinase	Phosphorylation	Activation	[32],[33]
PKGIα	Kinase	Cysteine oxidation	Activation	[34]
Src	Kinase	Cysteine oxidation	Inactivation	[35]
FOXO	Transcription factor	Acetylation, Phosphorylation, Ubiquitination	Activation	[36]
HIF1	Transcription factor	Deubiquitination	Enhanced DNA binding	[37]
P53	Transcription factor	Cysteine oxidation, Phosphorylation	Enhanced DNA binding	[38]
NF-κB (p50)	Transcription factor	Cysteine oxidation	Reduced DNA binding	[39]
NF-κB (p65)	Transcription factor	Phosphorylation	Enhanced DNA binding	[40]
NF-κB/RelA	Transcription factor	Phosphorylation	Reduced DNA binding	[41]
Nrf2/Keap1	Transcription factor	Deubiquitination	Nrf2-ARE binding; activation	[42],[43]
PTPs	Phosphatase	Cysteine oxidation	Inactivation	[44]
PTP1B	Phosphatase	Cysteine oxidation	Inactivation	[45]
PTEN	Phosphatase	Cysteine oxidation	Inactivation	[46]

2.1. Redox signaling

Cysteine oxidation/sulfenylation (-SOH) mediated activity change of signaling effectors is a well conserved redox responsive mechanism. It is direct and reversible sensing of redox potential within cellular milieu. It also provides rapid response to adjust the effect of growth signaling in order to cope with unfavorable environment. Notably, a number of phosphorylation consensus motifs are adjacent to cysteine residues that are subject to oxidation. For example, oxidized cysteines interfere with AKT-dependent phosphorylation of its substrate FOXO3 and override inhibitory PI3K/AKT signaling under oxidative stress [47]. As a result, FOXO3 is activated even in the presence of activated PI3K/AKT signaling in NSPCs. Similarly, reversible cysteine oxidation within the activating segment of Aurora kinase A interferes with adjacent threonine phosphorylation [48]. This mechanism is predicted to be dominant and evolutionarily conserved. Growing list of proteins that are subject to cysteine oxidation-mediated signaling is shown in Table 2. Below, we discuss a few examples of signaling effectors responsive to redox stress in NSPCs.

Table 2.

The list of molecules regulated by reversible cysteine oxidation

Molecule	Residue	Cell/Tissue type	Family/Class	Function	Reference
Bcl-2	C158 C229	human lung epithelial cells (H460)	Adaptor protein	Interfering interaction with ERK; apoptosis	[49]
DJ1	C106	H1299 HUVEC	Adaptor protein	Induced interaction with p53	[50, 51]
hSESN2	C125	HEK293	Adaptor protein	mTORC1 activation	[52]
STING	C148	HEK293, Murine BMDM	Adaptor protein	Inhibition STING dimerization	[53]
Calbindin D28K	C187 C219 C257		Calcium binding protein	Activation of myo-inositol monophosphatase, Reduction of Ca2+-binding	[54]
Recoverin	C39		Calcium binding protein	Reduction of Ca2+-binding	[55]
DLDH	Unknown	Rat brain	Dehydrogenase	Inactivation	[56]
GAPDH	C149	Rat ventricular myocyte	Dehydrogenase	Inhibition of glycolysis	[57]
A20	C103		Deubiquitinase	Reduced enzyme activity	[58]
USP1	Unknown	U2OS,T98	Deubiquitinase	Reduced enzyme activity	[59]
MGL	C201 C208	Rat primary cortical neurons	Hydrolase	Reduced enzyme activity	[60]
SGFH	C60	Saccharomyces cerevisiae	Hydrolase	Reduced enzyme activity	[61]
AKT2	C124	NIH-3T3	Kinase	Enhanced kinase activity	[62]
ASK1	C250	HEK293	Kinase	T838 phosphorylation; JNK activation	[63]
ATM	C2991	Human primary fibroblasts (GM08399 B)	Kinase	Form S-S dimer, Activate downstream signaling	[24]
Aurora A	C290	HeLa	Kinase	Reduced kinase activity	[48]
IKK0α			Kinase	Reduced kinase activity	[28]
JNK2	C222	Human articular chondrocytes	Kinase	Enhanced kinase activity	[64]
MEKK1	C1238		Kinase	Inhibition of ATP-binding site	[65]
Rheb	unknown	HEK293	Kinase	Activation	[31]
p56lck	unknown	Peripheral blood T lymphocytes (PBL blasts)	Kinase	Enhanced kinase activity	[66]
Pin1	C113	Human brain SH-SY5Y	Kinase	Reduced kinase activity	[67], [68]
PKA	C358	HeLa	Kinase	Reduced kinase activity	[48]
PKIG1a	C42	Mouse cardiomyocytes	Kinase	Reduced kinase activity	[34]
Src	C277 C185	NCI-H292	Kinase	Y416, Y527 phosphorylation; activation	[69], [35]
Aldose Reductase	C298 C303	Rat heart	Oxidoreductase	Reduced enzyme activity	[70]
MsrA	C72		Oxidoreductase	Reduced enzyme activity	[71]
Prx-1	C52 C83 C173		Oxidoreductase	Eliminates hydrogen peroxide	[72]
PTEN	C124	NIH-3T3, HeLa	Phosphatase	Suppressed enzyme activity	[73], [46]
PTPσ	C1589		Phosphatase	Reduced enzyme activity	[74]
PTP1B	C215	A431 (human epidermoid carcinoma)	Phosphatase	Enzyme inactivation	[45], [75],[76]
SHP-1/2	C455 C459	A20 (Murine B-Cell lymphoma)	Phosphatase	Enzyme inactivation	[77], [78]
Cathepsin K	C25		Protease	Reduced maturation of precathepsin K	[79]
EGFR	C797	A431	Receptor kinase	Enhanced kinase activity	[80]
FGFR1	C488		Receptor kinase	Enhanced kinase activity	[69]
AP-1			Transcription Factor	Decreased DNA binding	[81]
FOXO3	C31	NSPC	Transcription Factor	Inhibit T32 phosphorylation; activation	[47]
FOXO4	C477	HEK293	Transcription Factor	Acetylated by p300; inactivation	[82]
NF-κB (p50)	C62		Transcription factor	Decreased DNA binding	[83]
P53	C124 C141 C182		Transcription factor	Enhanced DNA binding	[38]
Yap1	C303 C598		Transcription factor	Activation	[84]
hBCATm	C315		Transferase	Reduced enzyme activity	[85]
MST	C247		Transferase	Reduced enzyme activity	[86]
MTAP	C136 C223	HepG2	Transferase	Reduced enzyme activity	[87]
PRMT1	C101 C208		Transferase	Reduced enzyme activity	[88]

ATM.

Ataxia telangiectasia (AT) is an autosomal recessive genetic disorder characterized by neurodegeneration with progressive ataxia, variable immunodeficiency, premature aging and an increased disposition to cancer. Clinically, A-T patients showed defective antioxidative capacity [89]. ROS oxidize the ataxia telangiectasia mutated (ATM) in its active disulfide-crosslinked dimer form, which activates the DNA repair pathway. ATM-lacking cells failed to activate the DNA repair pathway and were hypersensitive to DNA damage under oxidative stress. Studies with ATM oxidation indicate ATM to be a redox sensor in the cells [24]; [90]; [91]. ATM is abundant in NSPCs and is downregulated during differentiation. ATM-lacking NSPCs cannot differentiate into neurons or astrocytes and have shown abnormal proliferation and genomic instability [92].

PI3K/AKT/mTOR signaling.

PI3K/AKT signaling regulates ROS-dependent induction of neurogenesis. Also, promotion of the mammalian target of rapamycin (mTOR) pathway by ROS alters NSPC differentiation and migration, dendrite development, and neuronal maturation. Disruption of the mTOR pathway results in a host of neurodevelopmental disorders [93].

Evidence indicates the importance of ROS and PI3K/AKT signaling in NSPC differentiation and neuronal maturation. It also suggests that an enhanced oxidative cellular status and increased PI3K/AKT signaling in NSPCs lead to premature neurogenesis and prevention of excessive proliferation [94]. However, it is important to note that a different study found that ROS-mediated enhancements in the self-renewal and neurogenesis of NSPCs are dependent on PI3K/AKT signaling. ROS activates the PI3K/AKT/mTOR pathway via oxidative inactivation of the PTEN protein, a negative regulator of PI3K signaling. PTEN counteracts the kinase function of PI3K, subsequently inhibiting AKT activation. This PI3K-dependent activation of AKT is crucial for the growth and proliferation of adult hippocampal stem/progenitor cells [95]. Also, H₂O₂ deactivates PTEN, which enhances AKT signaling and maintains physiological signaling and proliferation of adult hippocampal stem/progenitor cells [96]. Thus, high ROS status of NSPCs seem to be required in order to maintain adequate levels of PI3K signaling and in turn maintain self-renewal and neurogenesis [14].

Wnt.

Elevated ROS levels enhance activation of Wnt/β-catenin signaling, which regulates the efficiency of neural differentiation. Evidence indicates that ROS production precedes Wnt/β-catenin activation, and that mitochondrial ROS metabolism is involved in regulating early events of Wnt/β-catenin signaling transduction. Delaying the onset of Wnt/β-catenin activation will lead to impaired neuronal differentiation [97]. The activation of Wnt/β-catenin signaling can mediate neuroprotection and improve neurological function during oxidative stress via Nrf2/PI3K/AKT-Wnt/Fzd-1/β-catenin cooperation and as part of a feedback loop regulating cellular homeostasis [98].

Type-I Interferon response.

The type-I interferon response is an innate immune response. An increase in type-I interferon signaling is found to be correlated with increased oxidative stress, as well as aging in the brain [99]; [47]. ROS signaling activates FOXO, which triggers the activation of the type-I interferon pathway. This increased type-I interferon response under oxidative stress suppresses the proliferation of NSPCs and reduces neuronal differentiation [47]. Type-I interferon response also activates cellular autophagy [100] and may serve as a protective response against oxidative stress.

2.2. Redox responsive transcription factors

A primary defense mechanism to oxidative stress is the antioxidant response, which combats noxious effects of oxidative insults [98]. Antioxidants rescue cells from oxidative stress and preserve redox balance by promoting defenses to neutralize ROS and regulate transcription [101]. Anti-oxidative stress responsive transcription factors and defensive gene expression program, therefore, are critical for maintaining NSPC reserves and their function. PR domain containing 16 (PRDM16), a histone-lysine N-methyltransferase, is required for NSPC maintenance and regulation of neural progenitor behavior [102]. PRDM16 depletion results in an elevation of oxidative stress levels and drives the expression of oxidative stress-responsive genes in cortical radial glial progenitors (RPGs). Removal of PRDM16 is also found to alter neocortical organization and expand RPG proliferation in the embryonic stage (E13.5) [103]. In addition to PRDM16-mediated fitness keeping constitutive transcription, below are several transcription factors acutely responsive to redox stress.

NRF2.

Nuclear factor erythroid 2-related factor 2 (Nrf2) is the master regulator of the cellular defense against oxidative stress [98]. Nrf2 activity is controlled by Kelch-like erythroid cell-derived protein with CNC homology (ECH)-associated protein 1 (Keap1) which serves as a cysteine-based intracellular sensor for oxidative stress [104]; [105]. Nrf2 regulates antioxidant gene expression through binding to the antioxidant-responsive element (ARE) (TGA[C/T]NNGC) and modulates mitochondrial biogenesis and function. By improving mitochondrial function, Nrf2 is able to diminish intracellular ROS overproduction [106].

Nrf2 is crucial for NSPC function. Nrf2 knockout mice showed impaired NSPC stemness and neuronal differentiation [107]. Aging downregulated the Nrf2-Hmox adaptive response and thus impaired neuronal differentiation of NSPCs in Parkinson’s disease (PD) mouse model [108]. During aging, a diminishing Nrf2 expression compromises the regenerative potential of NSPCs and results in a specific temporal pattern of decline in neurogenesis [109]. Overexpression of Nrf2 in ‘old’ NSPCs rendered them similar to ‘young’ NSPCs, showing increased survival and regneration. This implies that Nrf2 is a significant influencer in NSPC survival and function during aging [110].

HIF-1α.

Hypoxia leads to an upregulation and stabilization in hypoxia inducible factor-1α (HIF-1α) levels [111]. This is due to a paradoxical increase of oxidative stress under hypoxia. Electrons leak from the mitochondrial electron transport chain generating ROS under low oxygen tension and increasing HIF-1α [112]. NSPCs are exposed to a hypoxic environment during embryonic development. Thus, hypoxia signaling plays an important role in regulating NSPC maintenance and differentiation. Under hypoxic conditions, HIF-1α promotes signaling pathways that promote NSPC self-renewal and inhibits pathways that promote differentiation [113]. HIF-1α is also crucial to maintaining NSPCs as well as the vascularity of the niche [114]. Similarly, genetic inactivation of HIF-1α reduced adult hippocampal NSPCs and impaired neurogenesis [115].

TP53.

Loss of function of the tumor suppressor gene p53 results in elevated ROS, leading to early neurogenesis [116]. Restoration in function of p53 and anti-oxidative treatments partially reverse this phenotype, suggesting that p53-mediated regulation of physiological ROS levels may be required for the appropriate timing of neurogenesis. p53 has also been identified to play a role in neuronal fate via mediation of ROS levels [117]. The mitochondrial translocation of p53 is associated with a decrease in mitochondrial ROS levels and a shift in differentiation toward neurons, rather than astrocytes.

NFκB.

Nuclear factor-κB (NFκB) signaling is a key regulator for multiple niche factors that are involved in regulating different stages or phases of neurogenesis. The activation of NFκB is important in regulating self-renewal and early differentiation, as well as apoptosis of NSPCs [118]. NFκB is also a redox-sensitive transcription factor activated by ROS [119]. NFκB plays a protective role under oxidative stress by suppressing ROS accumulation and activating antioxidant targets [120]. Conversely, depending on phase and context, ROS can have bimodal effects and can either activate or suppress NFκB signaling. Oxidative stress activates the NFκB pathway in the early phase and suppresses NFκB activation in the late phase [120].

FOXO.

Forkhead box O (FOXO) transcription factors have protective effects against aging-progressive axonal degeneration through the suppression of mTORC1-induced proteotoxic stress to restore autophagy capacity and reduce ROS [121]. As a critical defense mechanism against oxidative stress FOXOs emerged as a major determinant of neural cell fate [122].

FOXOs are important for the maintenance of NSPC homeostasis and have protective effects against ROS [15]. FOXO1 regulates neurogenesis and interacts with the Notch pathway to control NSPC fate [123]. FOXO3 regulates glucose or carbon metabolism in a manner that supports redox balance and the neurogenic potential of NSPCs [124]. Under oxidative stress, FOXO3 promotes upregulation of glycine-N-methyltransferase (GNMT), a methyltransferase for glycine, which leads to s-adenosylmethionine (SAM) depletion, activation of type I interferon response, and suppression of differentiation, sequentially [47]. These FOXO-mediated oxidative stress responses protect NSPCs from premature activation and maintain their quiescence.

Notably, direct protein modification by cysteine sulfenylation was proposed to be an oxidative stress sensing mechanism of FOXO3 [47]. Cysteine sulfenylation adjacent to inhibitory AKT phosphorylation site of FOXO3 circumvents PI3K-AKT-mediated inactivation signal under oxidative stress (Figure 2). This mechanism is also highly conserved in a number of redox signaling cascades as discussed above.

Figure 2. Integration of redox sensors and effectors.

Receptor tyrosine kinases (RTK) produces ROS along with regulating PI3K/PTEN/AKT. AKT phosphorylates and inhibits nuclear translocation and activation of FOXO. Metabolic stress-induced ROS activates LKB/AMPK, which, in turn, activates autophagy and inhibits mTORC1 activity. Oxidative stress-activated SIRT and AMPK increase mitochondria biogenesis through PGC1α. ROS stabilize Nrf2 or HIF1α by dissociation of Keap1 and VHL, respectively. Cysteine oxidation among other modifications plays an important role in the response.

2.3. Redox stress-induced epigenetic alterations

Chromatin remodeling is an energy-demanding cellular process, and therefore requires balance with cellular metabolism and availability of metabolites. Several metabolic pathways produce intermediates that are involved in modifying the epigenetic landscape, and subsequently regulating NSPC fate. Oxidative stress is demonstrated to cause remodeling of chromatin and epigenetic reprogramming [125].

One-carbon metabolism drives epigenetic changes via two pathways: the folate pathway, which is involved in DNA synthesis, and the methionine pathway, which is involved in methylation reactions [126]. This metabolic pathway is involved in regulating NSPC self-renewal and differentiation. A recent study showed chromatin serves as methyl group sinks when phospholipid methylation is impaired [127]. Excess s-adenosylmethionine (SAM) is consumed by histone methylation reactions promoting trimethylation of histone H3 on Lys4 (H3K4me3), H3K36me3 and H3K79me3. These histone modifications could contribute to the altered gene expression, but they were not directly correlated with stable changes in transcription. Instead, histone methylation reaction facilitates SAM to s-adenosylhomocysteine (SAH) conversion, which then feeds into the transsulfuration reaction to boost anti-oxidant glutathione synthesis. Whether altered histone modification contributes to gene expression changes leading to NSPC fate change has not been reported.

The Eph:ephrin cell-cell communicating signaling regulates one-carbon metabolism to control NSPC differentiation. Ephrin B1 stimulation of NSPCs alters the methylation state of H3K4 by suppressing dihydrofolate reductase expression, a key enzyme for cellular proliferation. Eph forward signaling decreases H3K4 methylation on progenitor genes (Sox2, Nes) to promote differentiation in NSPCs [128]. Interestingly, this epigenetic reprogramming is long-lasting and inherited that NSPCs are locked in a differentiation-prone state. Given that dihydrofolate reductase engages NADPH as a cofactor, its action is likely to be sensitive to intracellular oxidative stress. H3K4me3 peak breadth has been linked to the cellular identity and transcriptional consistency in NSPCs [129]. How elevated redox stress may affect the H3K4me3 landscape and fate of NSPCs and their fate remains to be defined.

SAM, the principal methyl donor in cellular methylation reactions, is exhausted under oxidative stress [125]. SAM metabolism is required for the preservation of proliferative signaling in NSPCs. Oxidative stress is shown to suppress the neurogenic potential of NSPCs by activating type-I interferon response through reduction of intracellular SAM availability [47]. Depletion of SAM during oxidative stress response resulted in decreased nuclear lamin methylation which compromised nuclear envelope integrity. Nuclear lamina integrity is required for the proper 3D organization of nuclear chromatin [130]. Although not directly, redox stress is likely to have a profound role in both epigenetic and genomic reorganization in NSPCs.

The glutamine metabolism is essential for NSPC growth and long-term maintenance. It is a precursor for glutamate, a key building block of glutathione and therefore protects NSPCs from oxidative stress [124]. Glutamine also feeds into α-ketoglutarate, which is a key cofactor for histone and DNA demethylases. It is required for the removal of repressive histone modifications and DNA methylation in NSPCs to promote multipotency [131]. The metabolic reprogramming from aerobic glycolysis to the TCA cycle is associated with neuronal differentiation of NSPCs. Several TCA intermediates, including α-ketoglutarate, NADH, and FADH₂, are involved in the epigenetic regulation of transcription [132].

Lipid metabolism is involved with promoting histone acetylation through acetyl-CoA and expression of growth-related genes to drive cellular growth [133]. This metabolic pathway plays a significant role in the interaction between adult NSPCs and their niche, and promotes NSPC proliferation and neurogenesis [132]; [134]. Altered histone acetylation and associated gene expression changes under oxidative stress have been reported [125]. Previously, the role of acetyl-CoA generated by acetyl-CoA synthetase 2 (ACSS2) in hippocampal neuronal differentiation has been demonstrated [135]. ACSS2 is localized to the nucleus and functions as a chromatin-bound transcriptional coactivator that stimulates histone acetylation and gene expression relevant to the plasticity of neurons. Similarly, cellular acetyl-CoA metabolism and neuronal differentiation connection has been reported. TP53-inducible glycolysis and apoptosis regulator (TIGAR) is an endogenous inhibitor of glycolysis and is upregulated during neuronal differentiation [136]. Depletion of TIGAR reduced acetyl-CoA and H3K9 acetylation at the promoters of neurogenic genes resulting in decreased neuronal differentiation. Acetate, a precursor of acetyl-CoA, restored H3K9 acetylation and NSPC differentiation of TIGAR-depleted cells. Given that TIGAR plays a role in oxidative stress-induced metabolic reprogramming [137], it is likely to regulate NSPC differentiation through a histone acetylation-based epigenetic mechanism.

Oxidative stress has been found to alter metabolites and disrupt the epigenetic state of the cell [138]. Changes in the redox state was shown to regulate the fate of NSPCs via histone deacetylase SIRT1. Under oxidative stress conditions, SIRT1 is upregulated through increased NAD+ to NADH ratio [139]. Oxidative stress-activated SIRT1 suppresses adult hippocampal NSPC self-renewal [140] and inhibits the pro-neuronal MASH1 transcription factor expression, diverting the fate of NSPCs from neurogenesis towards the astroglial lineage [141].

2.4. Redox stress-activated autophagy

Recent studies have begun to look at the role of autophagy in the regulation of neurogenesis in the adult brain. The available data indicate the autophagy pathway regulates processes including maintenance and activation of quiescent NSPCs [142]. FOXO transcription factors play a critical role in maintaining physiological autophagic flux to regulate neurogenesis [143]. Autophagy is vital for cellular homeostasis in removing protein aggregates and dysfunctional organelles for physiological neuronal development.

Evidence demonstrates a vital role for autophagy in neuronal differentiation. It acts as a response-survival mechanism to reduce oxidative stress and regulate synaptogenesis that is associated with this process [144]. The downregulation of pro-autophagic proteins Ambra1 and Beclin 1 in adult NSPCs results in a decrease in cell proliferation and enhanced sensitivity to DNA damage-induced cell death [145]. The findings from this study suggest that Ambra1- and Beclin 1-mediated autophagy is involved in adult neurogenesis by controlling the survival of NSPCs.

Increasing evidence suggest the crosstalk between ROS and autophagy may play a determinant role in the regulation of neuronal homeostasis. Under conditions of oxidative stress, mitochondrial ROS induce autophagy [146]. This activation of autophagy is key in the cellular response to oxidative stress in order to dispose of defective components and prevent further damage to cells. Excessive ROS production not only impairs cellular macromolecules, but can also cause damage to antioxidant enzymes and nonenzymatic oxidants [147]. The autophagic process is thus critical in maintaining homeostasis by serving as a buffer system to control ROS levels in cells and inhibit inappropriate activation of ROS in response to neuronal damage [148]

2.5. Metabolic flexibility against oxidative stress

The coordination of metabolic adaptation with cellular signaling and epigenetic regulation supports divergent NSPC cell fate [149]. Distinct metabolic processes and their involvement in the regulation of NSPC fate have been studied to provide insight into the regenerative potential of the brain [5]. For example, subventricular zone NSPCs mostly rely on fatty acid oxidation to produce energy and support their neurogenic activity [150]. Similarly, hippocampal NSPCs may operate on beta-oxidation and glycolysis as the major metabolic activity as inferred from single cell transcriptome analysis [151]. NSPCs also rely on the pentose phosphate pathway (PPP) and thus are sensitive to its perturbation [124]. Oxidative PPP forms reduced nicotinamide adenine dinucleotide phosphate (NADPH), which is important for the suppression of cellular oxidative stress [152]. Oxidative stress redirects glucose into the PPP by a feedback regulatory loop and generates NADPH, suppressing an elevation of ROS [153]; [154]. In addition, glutamine flux regulates mTOR activity, translation, and autophagy to coordinate cellular growth and proliferation [155]. Glutamine metabolism also contributes to the control of redox balance by increasing the NADPH/NADP(+) ratio to suppress oxidative stress in NSPCs [124]; [156].

Mitochondrial dynamics is central to metabolic adaptation under oxidative stress. As a central organelle generating intracellular ROS, mitochondrial function and morphology are closely associated with NSPC fate [116]. The plasticity of the mitochondria enables it to undergo morphological and functional changes in response to cellular demands. This mitochondrial plasticity plays a crucial role in integrating signaling cues to regulate stem cell fates [157]. Changes in mitochondrial dynamics modulate NSPC fate decisions by driving a physiological ROS-mediated process that suppresses self-renewal and promotes differentiation [158]. Dysfunctional mitochondria due to mitochondrial DNA mutations impairs oxidative phosphorylation and accumulation of ROS, resulting in oxidative stress and altered redox signaling pathway activity. Unlike mitochondria-derived physiological ROS signaling, high levels of ROS produced from dysfunctional mitochondria impair adult neurogenesis [159]. Conversely, redox signaling through cysteine modifications regulate mitochondrial function and metabolism [160].

Studies reveal changes in mitochondrial morphology as NSPCs undergo neuronal differentiations Mitochondrial morphology changes regulate mitochondrial function of metabolism and ROS generation. Mitochondrial dynamics acts as an upstream regulator of physiological ROS to control NSPC self-renewal and fate decisions [158]. There is a demonstrated increase in vulnerability of mitochondrial mass and DNA to oxidative damage during NSPC differentiation. Maintaining mitochondrial DNA integrity is vital for the proper maturation of the mitochondria during neural differentiation [161]. The accumulation of mitochondrial damage from oxidative stress would result in NSPCs failing to properly differentiate into neurons.

3. Conclusion

The above body of evidence supports that cellular redox states regulate the balance between NSPC quiescence maintenance and activation in adult mammalian brains. The interplay of key redox sensors and effectors is depicted in Figure 2. In general, increased levels of intracellular ROS were linked to proliferation and differentiation of NSPCs [162]. A more recent study, however, revealed that quiescent NSPCs of the adult hippocampus unexpectedly show the highest levels of ROS [13]. This finding adds more complexity to the link between redox regulation and NSPC fate decision. While many studies addressed the causality of redox stress in NSPC fate changes the majority of observations are based on perturbative and correlative conditions. Further studies using real tine tracing of NSPC cell fate with the biosensor-based quantitation should aid better appreciation of the role of dynamic redox balance. The interplay between NSPC microenvironment and cellular autonomy also needs to be better appreciated. With a recent advent of experimental platforms including cerebral organoids and assembloids, induced vascularization and single cell genomics, it is possible to directly address how redox potential may control NSPC fate without perturbing heterotypic interaction with other cell types. In a recent translational study, a personalized cell-therapy strategy of implanting autologous induced pluripotent stem cells into a patient with PD demonstrated clinical improvement coinciding with reinnervation of the putamen by projections from dopaminergic neurons [163]. This successful proof of principle study further speaks for the importance of steering NSPC cell fate in vivo. A better understanding of underlying redox-based molecular mechanisms may contribute to the development of methods to optimally stimulate neuronal differentiation of NSPCs in patients with neurodegeneration and brain damages.

Highlights.

• NSPC is vulnerable to oxidative stresses due to the surrounding brain with high oxygen consumption.

• Redox-stress sensing and coping mechanisms balance NSPC self-renewal and differentiation.

• Direct cysteine modifications of signaling intermediates and transcription factors result in metabolic reprogramming, epigenetic alterations towards adaptive responses under increased redox potential.

Abbreviations

aaRS

Alanyl-tRNA synthetase

ACSS2

Acyl coenzyme A synthetase-2

AKT

Protein kinase B

AMPK

5’adenosine monophosphate-activated protein kinase

ASK1

Apoptosis signal-regulating kinase 1

ATM

Ataxia telangiectasia mutated

Bcl-2

B-cell lymphoma 2

DLDH

Dihydrolipoamide Dehydrogenase

EGFR

Epidermal growth factor receptor

ERK

Extracellular signal-regulated kinase

FADH₂

Flavin adenine dinucleotide

FGFR1

Fibroblast Growth Factor Receptor 1

FOXJ1

Forkhead box protein J1

FOXO

Forkhead box protein O

GAPDH

Glyceraldehyde 3-phosphate dehydrogenase

GNMT

Glycine N-methyltransferase

hBCATm

Human branched chain aminotransferase

HIF1

Hypoxia inducible factor I

hSESN2

Human sestrin-2

IKK

Inhibitor of nuclear factor-κB kinase

IQGAP

Ras Guanosine-5’-triphosphatase-activating-like protein

JNK

c-Jun N-terminal kinases

Keap1

Kelch-like erythroid cell-derived protein with CNC homology-associated protein 1

MASH1

mammalian achaete scute homolog-1

MEKK1

Mitogen-activated protein kinase kinase kinase-1

MGL

Monoacylglycerol lipase

MsrA

Methionine sulfoxide reductase A

MST

Macrophage-stimulating protein

MTAP

S-methyl-5’-thioadenosine phosphorylase

mTORC1

Mammalian target of rapamycin complex 1

NADPH

Nicotinamide adenine dinucleotide phosphate

NF-kB

Nuclear factor-kB

NOX

NADPH oxidase

Nrf2

Nuclear factor erythroid 2–related factor 2

NSPCs

Neural stem/progenitor cells

P38 MAPK

p38 mitogen-activated protein kinase

P56lck

p56 lymphocyte-specific protein kinase

PI3K

Phosphoinositide 3-Kinase

PKA

Protein kinase A

PKG1a

Protein kinase G type- 1alpha

PPP

Pentose phosphate pathway

PRDM16

PRDF1-RIZ (PR) domain containing 16

PRMT1

Protein arginine N-methyltransferase-1

Prx-1

Peroxiredoxin-1

PTEN

Phosphatase and tensin homolog

PTP

Protein tyrosine phosphatase

ROS

Reactive oxygen species

SAM

S-adenosylmethionine

SHP-1/2

Src homology 2 domain tyrosine phosphaoases-1/2

SIRT1

Sirtuin-1

STING

Stimulator of interferon genes

TCA

Tricarboxylic acid cycle

TIGAR

TP53 Induced Glycolysis Regulatory Phosphatase

TRPA

Tryptophan synthase, alpha subunit

USP1

Ubiquitin Specific Peptidase 1

VEGFR

Vascular endothelial growth factor