Sestrin2 as a gatekeeper of cellular homeostasis: Physiological effects for the regulation of hypoxia‐related diseases

Abstract Sestrin2 (SESN2) is a conserved stress‐inducible protein (also known as hypoxia‐inducible gene 95 (HI95)) that is induced under hypoxic conditions. SESN2 represses the production of reactive oxygen species (ROS) and provides cytoprotection against various noxious stimuli, including hypoxia, oxidative stress, endoplasmic reticulum (ER) stress and DNA damage. In recent years, the determination of the regulation and signalling mechanisms of SESN2 has increased our understanding of its role in the hypoxic response. SESN2 has well‐documented roles in hypoxia‐related diseases, making it a potential target for diagnosis and treatment. This review discusses the regulatory mechanisms of SESN2 and highlights the significance of SESN2 as a biomarker and therapeutic target in hypoxia‐related diseases, such as cancer, respiratory‐related diseases, cardiovascular diseases and cerebrovascular diseases.


| INTRODUC TI ON
Hypoxic injury is a complex pathophysiological process involving a variety of factors. Acute hypoxia can cause pulmonary arterial hypertension, decrease in myocardial contractility and arrhythmia. 1 Lack of oxygen and blood supply to the brain can cause hypoxicischaemic encephalopathy (HIE), which results in severe brain damage, and poses a significant threat to learning and memory functions. 2 Similarly, myocardial ischaemia injury occurs when the heart suffers from hypoxia and insufficient blood supply. Moreover, hypoxia is also a feature of the cancer microenvironment. Hypoxiainducible factor-1α (HIF-1α) is often overexpressed and accumulates in cancer cells, in which HIF-1α-mediated signalling is a crucial pathway that regulates the metabolism and growth of solid tumours. 3 Collectively, hypoxia causes abnormal changes in cell metabolism, function and morphology.
Sestrin2 (SESN2) belongs to the evolutionarily conserved Sestrin family and is also known as a hypoxia-induced 95 gene (HI95). 4 Many studies have demonstrated that SESN2 is a stress-inducible protein that responds to various insults, such as hypoxia, energy deficiency, genotoxic stress and oxidative stress. 5 A recent study indicated that SESN2 silencing could suppress mitochondrial biogenesis, reduce mitochondrial biological activity, and ultimately, aggravate hypoxic injury. 6 Animal model experiments indicated that overexpression of SESN2 could improve hypoxia-ischaemia injury. 5,7-13 SESN2 is regulated by HIF-1α and is strongly associated with the oncogenesis and prognosis of solid tumours. 3 Therefore, we believe that SESN2 might serve as a potential target to protect against hypoxia injury.
In this review, we summarize the latest advances regarding the compensatory mechanisms of SESN2 in hypoxia metabolism and the possible signalling pathways involved. First, we review the general background of SESN2. Subsequently, we discuss the common signalling pathways of SESN2. Finally, we consider the role of SESN2 in several hypoxia-related diseases. This review provides novel insights into the stress modulation of SESN2 against hypoxia.

| G ENER AL BACKG ROUND OF S E S N2
In 2002, Budanov et al 4 used cDNA microarray hybridization in an attempt to identify novel genes participating in cellular responses to prolonged hypoxia. They named a novel gene, Hi95 (SESN2), whose protein product shared significant homology with a p53-regulated GADD family member PA26. In 2008, they revealed that SESN2 provided an essential link between genotoxic stress, p53 and the mammalian target of rapamycin (mTOR) signalling pathway. 14 Their study demonstrated that SESN2, as a target gene of P53, could activate the AMP-responsive protein kinase (AMPK) and inhibit mTOR. Subsequently, studies have confirmed that SESN2 is a stress-inducible protein that responds to various insults, such as hypoxia, oxidative stress, endoplasmic reticulum (ER) stress and DNA damage. 8,12,15 In addition, SESN2 plays a crucial role in cancer, metabolic disorders, cardiovascular diseases and neurodegenerative disorders. 10,13,16,17 Therefore, it is imperative to understand the functions of SESN2 in the modulation of its associated pathophysiological mechanisms and determine how SESN2 regulates hypoxia-related diseases.

| Hypoxia
Sestrin2 (SESN2) was first isolated as a gene that was activated in human neuroblastoma cells under hypoxia and was identified as a p53-dependent gene. 4 Subsequent studies found that SESN2 could be activated by energy deprivation secondary to prolonged hypoxia. 18 HIF-1 is a primary transcriptional regulator of cellular responses to hypoxia and has been shown to increase the expression of SESN2 in mouse epithelial tracheal cells exposed to oxidative stress. 19 When the environment of a cell is oxygenated, HIF-1α can be degraded rapidly by prolyl hydroxylases (PHDs). Under hypoxic conditions, the activity of PHDs is impaired, which results in stabilization of HIF-1α. 20 Seo et al 21  Therefore, SESN2-AMPK signal pathway increases the degradation of HIF-1α by regulating the activity of PHDs. Besides, HIF-1α is often overexpressed and accumulated in cancer cells, regulating the growth of solid tumours, such as pancreatic adenocarcinoma (PDAC or PAAD). 3 Using a gene expression profiling interactive analysis (GEPIA) data set (http://gepia.cance r-pku.cn/), we found that the expression levels of HIF1A and SESN2 were higher in PAAD than in normal samples ( Figure 1A,B), intriguingly, HIF1A was positively associated with SESN2 ( Figure 2). Since HIF-1α is mainly regulated post-transcriptionally, the positive associations between HIF-1α and SESN2 may not be important. Furthermore, Shi et al 11 studied the level of SESN2 expression in both severe and moderate hypoxicischaemic (HI) rat models. The data suggested that SESN2 was activated in severe HI, but not in moderate HI. Therefore, hypoxia can induce SESN2 expression. Although the mechanisms of SESN2 induction under hypoxia remained unclear, we speculated the SESN2 and HIF-1α interact with each other to regulate cellular metabolism.  21,32 The occurrence of cancers is associated with significant downregulation of SESN2. 21,33 Therefore, SESN2 is considered a vital factor in the removal of ROS and oxidative stress regulation.

| Endoplasmic reticulum stress
The accumulation of misfolded or unfolded proteins is known as ER stress. 34 Increasing evidence highlights the significant impact of ER stress on maintaining cellular homeostasis. 35 It was demonstrated that SESN2 expression was increased via the protein kinase R-like endoplasmic reticulum kinase (PERK), eukaryotic initiation factor-2 alpha (eIF2α)/activating transcription factor-4 (ATF4)-dependent pathway. 36 The unfolded protein response (UPR) is significant in oxidative stress and inflammatory responses in cancer. Ro

| Downstream effectors
3.2.1 | The AMPK/mTORC1 pathway AMP-responsive protein kinase (AMPK), which is regulated by the AMP to ATP ratio, is considered a cellular energy sensor and plays a critical role in energy metabolism. 45 The mechanisms underlying AMPK activation include hypoxia, glucose deprivation liver kinase B1 (LKB1) and SESN2. 46 mTOR is present in two distinct complexes, mTORC1 and mTORC2. It was reported that inhibitors of mTOR might be available to treat cancer, cardiovascular disease, autoimmunity F I G U R E 2 The correlation between HIF1A and SESN2 in PAAD HIF1A was positively associated with SESN2 (R = 0.56, P < 0.01) and metabolic disorders. 47 AMPK, as a negative regulator of mTOR signalling, phosphorylates and inactivates mTORC1.
It is widely accepted that SESN2 inhibits mTORC1 through the activation of AMPK. 14 GTPases to control mTORC1 signalling. In addition, accumulating evidence showed that the AMPK/mTORC1 signalling pathway was significantly associated with the role of SESN2 in genotoxic stress, 14 ROS elimination 50 and autophagy. 51 As previously described, SESN2, as a stress-inducible protein, plays a crucial role in cell homeostasis and metabolism through the AMPK/mTORC1 pathway (Figure 3), and further study of this pathway will contribute to our understanding of the regulation of cellular energy metabolism. pathway. 27 It was reported that the KEAP1/NRF2 signalling system plays a crucial role in a range of diseases, including inflammatory diseases, 55 stroke 56 and cardiovascular diseases. 57 In summary, SESN2 exerts its antioxidant defense effects via the elimination of ROS accumulation through KEAP1/NRF2 signalling activation.

| Autophagy
Autophagy is a cellular defense mechanism that is important for the maintenance of cellular homeostasis. 58 As described earlier, SESN2 regulates the AMPK/mTORC1 signalling pathway, and mTORC1 is a major regulator of autophagy. 58,59 In the presence of nutrients, mTORC1 is activated to inhibit the Unc-51-like protein kinase 1 (ULK1) complex and autophagy. 60 64 Taken together, these studies suggest that SESN2 plays an integral part in autophagy; however, further research is required to determine its beneficial effects and to develop ways to inhibit its adverse effects.

| S E S TIN2 AND D IS E A S E S
An increasing number of studies have confirmed the stress response impact of SESN2 on hypoxia metabolism; however, SESN2 behaves differently in various diseases. Here, we present an overview of SESN2 in hypoxia-related diseases to provide a reference for their subsequent treatment (Figure 3).

| Cancer
The occurrence of cancer is strongly associated with DNA damage, gene mutation, oxidative stress, metabolic dysregulation and inflammation. 65 Recent research indicated that SESN2 plays a crucial role in many kinds of cancer cells. Most cancer cells show enhanced survival in the hypoxic tumour microenvironment via HIF-1 overexpression. Several studies indicated that SESN2 levels were significantly decreased in colon adenocarcinoma tissues, and low expression of SESN2 promoted colon tumour growth. 16 In addition, high expression of SESN2 decreased murine colon tumour cell growth both in vitro and in vivo. 63,66,67 We compared the mRNA levels of SESN2 in colon tumours with those in normal samples using the ONCOMINE (https://www.oncom ine.org/) databases, which showed that the mRNA expression of SESN2 was significantly down regulated in colon tumours (Figure 4).
The mechanism by which SESN2 regulates tumour growth was investigated by Ro et al, 16 who proved that SESN2 was an essential mediator of p53's control over mTORC1 signalling and tumour cell growth. SESN2 inhibited colonic cell growth by suppressing both mTORC1 and ER stress. Accumulating evidence demonstrates that SESN2 can inhibit tumour growth and overcome antibiotic resistance, such as in prostate cancer, 68 neuroblastoma 59 and breast cancer. 69 In non-small cell lung cancer, high expression of SESN2 was associated with prolonged overall survival compared with that of patients with low SESN2 expression. 33 73 On the one hand, early-stage tumour cytogenesis is associated with decreased autophagy levels. On the

F I G U R E 4
The transcription levels of SESN2 in different types of cancers (ONCOMINE dataset, https://www.oncom ine. org/). Compared with that in normal tissue, SESN2 expression was downregulated in colorectal cancer, leukemia, and lymphoma other hand, during cancer maintenance, autophagy activity is upregulated. 74 We speculated that the dual effects of SESN2 in cancer regulation were related to its mediated autophagy. Moreover, HIF-1α/Bcl-2 and adenovirus E1B 19 kD-interacting 3(BNIP3)mediated mitochondrial autophagy plays a crucial role in solid tumours. 75 However, there is no research concerning the relationship between SESN2 and the HIF-1α/BNIP3 pathway.
In view of the dual role of SESN2 in cancer, further long-term studies are required.

| Respiratory related diseases
Obstructive sleep apnea (OSA), which exhibits intermittent hypoxia, could lead to arterial hypertension, stroke and other complications.
These complications are associated with the triggering of oxidative stress. 76 Bai et al 77 demonstrated that the level of urinary SESN2 in patients with OSA was significantly higher than that of the control group and increased with the severity of OSA. Thus, the level of urinary SESN2 might be a biomarker of OSA severity. Moreover, it was reported that oxidative stress increased in asthma and caused airway inflammation and airway remodelling. 78 Kang et al 79 demonstrated that patients with asthma had a significantly higher plasma SESN2 level than the control group. Although the underlying mechanism for the significant upregulation of SESN2 has not been determined in patients with OSA and asthma, SESN2 might be a promising biomarker for OSA and asthma. Further research should be conducted to clarify its specificity and sensitivity.

| Cerebral ischaemic diseases
Hypoxic-ischaemic encephalopathy (HIE) is the leading cause of morbidity and mortality in infants, and remains the primary cause of perinatal brain injury, resulting in varying degrees of disability. 80,81 It was reported that activation of the NRF2 and AMPK pathways were important in the treatment of cerebral ischaemia. 12,13 Wang et al 13 suggested that SESN2 promotes cerebral angiogenesis after ischaemia through the NRF2/HO-1 pathway. Furthermore, it was reported that SESN2 overexpression decreased the brain infarct volume and diminished neuronal injury. Shi et al 12 proved that SESN2 had a substantial neuroprotective effects after HIE via the AMPK/ mTOR pathway. Stabilization of the blood-brain barrier plays a crucial role after HI, and recombinant human (rh)-sestrin2 attenuated blood-brain barrier permeability and alleviated brain infarct and oedema by upregulating endogenous SESN2 levels. 11  Cerebral ischaemia/reperfusion injury (I/R) is a very complex pathophysiological process, which involves an overload of intracellular calcium (Ca), lipid peroxidation, oxygen free radical damage, apoptosis gene activation and inflammatory cytokine damage. [82][83][84] SESN2 is closely associated with these processes, especially oxygen free radical damage. A study by Du et al 85 demonstrated that SESN2 alleviated oxygen-glucose deprivation and reoxygenation-induced apoptosis via the NRF2 pathway. Besides, SESN2 overexpression markedly decreased cerebral I/R injury by upregulating NRF2 in the nucleus. 86 Conversely, SESN2 silencing exacerbated cerebral I/R injury through the AMPK/PPARG coactivator 1 alpha (PGC1α) pathway. 5 In summary, SESN2 cannot only reduce HIE and IR damage, but also significantly improves neurological function by activating various signalling pathways.

| Ischaemic heart diseases
Coronary atherosclerotic heart diseases, such as myocardial infarction and ischaemic stroke, are major causes of morbidity and mortality worldwide. 87 Oxidative stress is a major player in cardiac pathophysiology; therefore, it was speculated that SESN2 would be protective against cardiomyopathies. With ageing, the level of SESN2 in the heart declined, leading to an impaired AMPK/PGC-1α signalling cascade and aggravated ischaemic insults. 88 Yang et al 89 found that SESN2 suppressed the activated macrophage-mediated inflammatory response in MI via the inhibition of mTORC1 signalling.
In the drosophila heart, loss of dSesn resulted in cardiac malfunction. 50 Similar reports suggested that SESN2 exerts heart-protective effects in ischaemic heart disease. 7,8 However, it was reported SESN2 concentrations were increased in patients with chronic heart failure (CHF), and correlated positively with the severity of CHF; indeed, augmented SESN2 levels increased the occurrence of major adverse cardiac events significantly, suggesting poor outcome in patients with CHF. 90 According to the above studies, we speculated that SESN2 levels increased in a compensatory manner in response to oxidative stress in ischaemic heart disease. It appeared that a protective effect occurred when the SESN2 concentration was sufficient to cope with oxidative stress; however, the increase was not sufficient to counter it, which might have led to a false impression.
Although reperfusion treatment of the ischaemic myocardium was established as a highly beneficial therapy for MI, more severe injury might occur after the onset of reperfusion, which is called ischaemia-reperfusion injury (IR injury). 91 It was reported that pyruvate dehydrogenase (PDH), which can modulate the interaction between SESN2 and LKB1, could regulate energy metabolism to alleviate the cardiac damage caused by I/R injury. 92 Quan et al 9 also proved that SESN2 mediated AMPK activation to alleviate I/R injury.
Taken together, SESN2 may represent a biomarker of the extent of CHF and a novel target for the amelioration of cardiovascular diseases.

| D ISCUSS I ON
Data from various animal experimental models and clinical research indicate that SESN2 is a promising target for the treatment of hypoxia-related diseases in humans. In this review, we briefly discussed the upstream and downstream regulators of SESN2 ( Figure 3). Regulated by P53 and HIF-1α, SESN2 is an important regulator of metabolism and oxidative stress. SESN2 can diminish ROS accumulation and activate autophagy, thus suppressing cancer, respiratory-related diseases, cardiovascular diseases and cerebrovascular diseases (Figure 3). Although the advantageous or disadvantageous roles of SESN2 appear in different diseases, SESN2 provides a novel therapeutic target for the prevention of hypoxiarelated diseases and metabolic disorders. Further research is needed to determine the specific underlying mechanisms by which SESN2 exerts its functions.

ACK N OWLED G EM ENTS
This work was supported by the National Natural Science Foundation of China (grant numbers 31971106 and 18CXZ244).

CO N FLI C T O F I NTE R E S T
The authors declare no conflict of interest.