Snapshot: implications for melatonin in endoplasmic reticulum homeostasis

Abstract

The endoplasmic reticulum (ER) is an important intracellular membranous organelle. Previous studies have demonstrated that the ER is responsible for protein folding and trafficking, lipid synthesis and the maintenance of calcium homeostasis. Interestingly, the morphology and structure of the ER were recently found to be important. Melatonin is a hormone that anticipates the daily onset of darkness in mammals, and it is well known that melatonin acts as an antioxidant by scavenging free radicals and increasing the activity of antioxidant enzymes in the body. Notably, the existing evidence demonstrates that melatonin is involved in ER homeostasis, particularly in the morphology of the ER, indicating a potential protective role of melatonin. This review discusses the existing knowledge regarding the implications for the involvement of melatonin in ER homeostasis.

Abbreviations

AD

Alzheimer's disease

ATF6

activating transcription factor 6

BIP

binding immunoglobulin protein

CCK‐8

cholecystokinin octapeptide

Cd

cadmium

CHOP

growth arrest‐ and DNA damage‐inducible genes 153

CsA

cyclosporine A

DM

diabetes mellitus

eIF2α

eukaryotic translation initiation factor 2α

ER

endoplasmic reticulum

H/I

hypoxia/ischaemia

HCC

hepatocellular carcinoma

Hsp70

heat shock protein 70

IP3

inositol trisphosphate

IRE1

inositol‐requiring protein 1

IRS

insulin receptor substrate

LD

12‐h light:12‐h dark

LL

continuous light

PERK

PRKR‐like ER kinase

RER

rough ER

RHDV

rabbit haemorrhagic disease virus

SER

smooth ER

UPR

unfolded protein response

XBP1

X‐box binding protein 1

α‐SMA

α‐smooth muscle actin

Tables of Links

TARGETS
Other protein targets a	Enzymes c
Bax	BIP
Bcl‐2	eIF2α
GPCRs b	IRE1
Melatonin receptors	JNK
	PERK
	SP1

LIGANDS
Cadmium
Cyclosporine A
IP3
Melatonin

These Tables list key protein targets and ligands in this article which are hyperlinked to corresponding entries in http://www.guidetopharmacology.org, the common portal for data from the IUPHAR/BPS Guide to PHARMACOLOGY (Southan et al., 2016) and are permanently archived in the Concise Guide to PHARMACOLOGY 2015/16 (^a,b,cAlexander et al., 2015a,b,c).

Introduction

Melatonin, known as the ‘hormone of darkness’, is important in both animals (Brzezinski, 1997; Tosches et al., 2014) and plants (Arnao and Hernandez‐Ruiz, 2014). In mammals, melatonin is primarily produced by the pineal gland, displays characteristic daily and seasonal patterns of secretion and plays an important role in sleep regulation (Tosches et al., 2014; Gandhi et al., 2015). Melatonin and the melatonin receptor have been demonstrated to be associated with major depression (Hickie and Rogers, 2011), jet lag (Waterhouse et al., 2007; Sack, 2010), multiple sclerosis (Farez et al., 2015), diabetes mellitus (DM) (McMullan et al., 2013), cancer (Dauchy et al., 2014; Sigurdardottir et al., 2015; Ma et al., 2016), fibrosis in multiple organs (Hu et al., 2016) and drug addiction (Feng et al., 2013) and might represent novel treatment targets. In particular, the administration of melatonin is able to normalize the increased pro‐inflammatory profile observed during metabolic syndrome (Cano Barquilla et al., 2014), atherosclerosis (Hu et al., 2013) and sepsis (Galley et al., 2014). Furthermore, the presence of numerous transmitters originating from various sources, particularly noradrenaline, has been reported to regulate the production and secretion of melatonin in the pineal gland (Simonneaux and Ribelayga, 2003; Gonzalez et al., 2012). The melatonin produced by the pineal gland further influences the rest of the body (Borjigin et al., 1999), and it has been reported that the reproductive system is controlled by the daily rhythm in melatonin production (Tamarkin et al., 1985).

The endoplasmic reticulum (ER) has a remarkably complex structure consisting of a single bilayer with a network of sheets and dynamic tubules (Westrate et al., 2015). Numerous genetic and environmental insults have been demonstrated to disturb the ultrastructure and volume of the ER, leading to a buildup of misfolded proteins in this organelle (a condition called ER stress) and initiation of the unfolded protein response (UPR) to remove the misfolded proteins (Walter and Ron, 2011; Brodsky, 2012; Hetz, 2013; Mehnert et al., 2014; Yang et al., 2015). An abnormal ER and ER stress are emerging as key contributors to a growing list of human diseases (Tabas and Ron, 2011; Walter and Ron, 2011; Oakes and Papa, 2015), including cancer (Mahoney et al., 2011; Clarke et al., 2014), neurological disease (Roussel et al., 2013; Hetz and Mollereau, 2014) and metabolic diseases (Fu et al., 2011; Arruda et al., 2014; Yang et al., 2015). Notably, there is close relationship between ER structure and function, and defects in ER structure are associated with diseases (Westrate et al., 2015). The structure and morphology of the ER can be regulated by various factors (Chen et al., 2012; English and Voeltz, 2013), and melatonin has attracted increasing attention.

This review focuses on the relationship between melatonin and the ER. Firstly, we briefly summarize the normal structure and functions of the ER. We then discuss the existing implications for melatonin in ER homeostasis, and the regulation of ER stress by melatonin in various pathologies is subsequently introduced. The information compiled in this review should help scientists understand the involvement of melatonin in ER homeostasis.

Summary of the ER

The ER is an important intracellular membranous organelle in eukaryotic organisms that consists of an interconnected network of flattened, membrane‐enclosed sacs or tubes known as cisternae (Hu et al., 2011; Smith et al., 2011). The ER is located throughout much of the cytoplasm, and the membranes of the ER are continuous with the plasma and nuclear membranes (Giordano et al., 2013; Chung et al., 2015; Westrate et al., 2015). A close association exists between the ER and the mitochondria (Rowland and Voeltz, 2012; Arruda et al., 2014), Golgi complex (Zanetti et al., 2012; He et al., 2013) and peroxisome (van der Zand et al., 2012; Tabak et al., 2013). The ER is found in most types of eukaryotic cells, including the most primitive Giardia (Soltys et al., 1996), and can be classified into two types, namely, rough ER (RER) and smooth ER (SER), depending on the presence of protein‐manufacturing ribosomes on the surface (Reid and Nicchitta, 2015). The RER is mainly associated with protein synthesis, and the SER is mainly responsible for lipid metabolism, carbohydrate metabolism and detoxification (Friedman and Voeltz, 2011). In general, the ER is responsible for protein folding and trafficking (Rowland and Voeltz, 2012; Zanetti et al., 2012; Reid and Nicchitta, 2015), lipid synthesis (Rowland and Voeltz, 2012; Prinz, 2014) and the maintenance of calcium homeostasis (Smith et al., 2011; Stutzmann and Mattson, 2011; Rowland and Voeltz, 2012).

The correct folding of newly synthesized proteins is made possible by the involvement of several ER chaperone proteins, including protein disulfide isomerase, the heat shock protein 70 (Hsp70) family member that binds immunoglobulin protein (BIP or glucose‐regulated protein 78), calnexin, calreticulin and members of the peptidylpropyl isomerase family. Only properly folded proteins are transported from the RER to the Golgi apparatus. Numerous environmental, physiological and pathological stressors, nutrient fluctuations and chemical triggers disturb the ER protein folding environment (Zaouali et al., 2010; Brown et al., 2014; Carloni et al., 2014; Chan et al., 2015; Garcia‐Marques et al., 2015); these factors cause protein misfolding, resulting in the accumulation of misfolded or unfolded proteins, a condition defined as ER stress (Wang and Kaufman, 2014).

The UPR is a collection of signalling pathways that have evolved for the maintenance of a productive ER protein folding environment (Wang and Kaufman, 2014). The UPR comprises three parallel signalling branches: PRKR‐like ER kinase (PERK)–eukaryotic translation initiation factor 2α (eIF2α), inositol‐requiring protein 1 (IRE1)–X‐box binding protein 1 (XBP1) and activating transcription factor 6 (ATF6) (Santos et al., 2014; Wang and Kaufman, 2014). Under non‐stress conditions, BIP binds to the domains of PERK, IRE1 and ATF6 to stabilize them and prevent their activation (Ozcan et al., 2004; Cnop et al., 2012). Stress can promote the separation of the BIP from PERK, IRE1 and ATF6 and the subsequent activation of these three molecules (Figure 1). Briefly, PERK autophosphorylation promotes the phosphorylation of eIF2α to strongly inhibit mRNA translation, shuts down global protein synthesis and induces an increase in activating transcription factor 4 (ATF4). IRE1 works as an endoribonuclease by slicing the mRNA of XBP1. ATF6 is processed by membrane‐bound transcription factor site‐1 (also known as site‐1 protease, SP1) and SP2 in the Golgi complex to generate cleaved ATF6. ATF4, XBP1 and cleaved ATF6 promote genes encoding ER chaperones. Taken together, these results indicate that the triggering of ER stress and activation of the UPR might be associated with a pro‐survival or pro‐apoptotic outcome via different mechanisms, which ultimately determine the fate of cells (Santos et al., 2014). BIP, PERK, eIF2α, ATF4, growth arrest‐ and DNA damage‐inducible genes 153 (GADD153 or CHOP, whose transcription is activated by ATF4; Harding et al., 2000; Palam et al., 2011; Wang and Kaufman, 2014), IRE1, XBP1, ATF6, BIP and glucose‐regulated protein 94 (GRP94) are often used as ER stress markers.

Figure 1.

ER stress and UPR signalling pathways. Unfolded proteins (UPRs) can promote the separation of the BIP from PERK, IRE1 and ATF6. PERK autophosphorylation then promotes the phosphorylation of eIF2α to strongly inhibit mRNA translation and shut down global protein synthesis; eIF2α also causes an increase in ATF4; IRE1 works as an endoribonuclease, slicing the mRNA of XBP1; and ATF6 is processed by SP1/2 proteins in the Golgi complex to cleave ATF6. ATF4, XBP1 and cleaved ATF6 promote the transcription of genes encoding ER chaperones. In the end, unfolded proteins are decreased, but ER chaperones are activated. SP1/2, site‐1 and site‐2 protease.

Melatonin and the ER in the pineal gland

Seasonal variation in the volume densities of the ER of at least some tissues is distinct and synchronized by changes in the photoperiod regulated by the pineal gland and the melatonin rhythm (Munoz et al., 2001). This finding indicates that melatonin has implications in ER homeostasis. As an important type of indoleamine, the synthesis of melatonin and associated key enzymes in the pineal gland may be closely associated with the state of the ER, particularly the RER (Kappers, 1978; Tan et al., 2015). Higher melatonin secretion appears to be partly related to the existence of developmental changes in the morphology and quantity of pinealocytes (the functional units of the pineal gland), which are characterized by a relatively large volume of RER (Redondo et al., 2003). Due to the diurnal rhythm of pineal melatonin production, the plasma melatonin concentrations are significantly higher at 02:00 h (at night) than at 09:00 h, 14:00 h (during the day) and 21:00 h (early night), but no significant differences in the RER are discernible (Lewczuk et al., 2004). Correlated with the well‐known nocturnal enhancement in the secretion of melatonin from the pineal gland, higher relative volumes of granular ER are found at night than during the daytime in pinealocytes (Karasek et al., 1990; Swietoslawski and Karasek, 1993). This finding indicates that the ER might be closely related to the secretion of melatonin by the pineal gland.

In the pineal gland of old shrews, the parenchyma undergoes alterations that mainly affect the ER cisternae, which show increased numbers of dense bodies, the formation of many concretions and a depletion of presumed secretory products (Dekar‐Madoui et al., 2012). These changes indicate that the gradual changes in the ER cisternae that occur during aging in Crocidura russula represent a reduced pinealocyte metabolism. Various stressors, including sleep deprivation and weaning of offspring, result in changes in the physiological function and morphological character of the ER, and these changes are reversed by melatonin supplementation. Thus, in goat kids subjected to premature weaning, melatonin might contribute to improvements in the histophysiological function of the ER (Redondo et al., 2010). The mean concentrations of melatonin in the plasma of weaned goat kids are significantly lower than those in non‐weaned offspring. A quantitative ultrastructural analysis of pinealocytes showed that the relative volume of the RER in pineal glands is significantly lower in weaned goat kids than in non‐weaned goat kids; however, treatment with melatonin significantly increased the RER volume in weaned kids (Redondo et al., 2010). Additionally, as a result of sleep deprivation, many pinealocytes exhibit dilation of the cisternae of the RER and SER, and these changes are attenuated by a single injection of melatonin (Lan et al., 2001). Collectively, these findings indicate that exogenous melatonin might promote a positive feedback in its pineal production through ER regulation.

Melatonin and ER function in target glands

Exogenously administered melatonin not only affects the ER morphology of the pineal gland but also regulates that of numerous organ systems, including the reproductive organs, the parathyroid gland and cancer cells. Melatonin directly or indirectly alters the ultrastructural appearance of mouse Leydig cells and influences their secretory activity by inhibiting their capacity to secrete steroids (Reiter et al., 2009). Melatonin treatment induces an obvious reduction in the volume of the SER, RER mitochondria and Golgi complex (Redins et al., 2002). Similarly, in adult Syrian hamsters, melatonin treatment alters the morphology of the seminal vesicles and ventral prostate, which presents conspicuous secretory granules and parallel arrays of granular ER with narrow cisternae (Chow and Pang, 1989). Furthermore, the cytoplasm of the coagulating glands and dorsal prostate of non‐melatonin‐treated animals are dominated by distended cisternae of granular ER and apical blebbing of endothelial cells. The administration of melatonin induces structural modifications, reflecting a loss of functional activities or even death of accessory sex gland secretory cells and thus confirming the suppressive effects of melatonin on the reproductive structures of rodents. Although the actions of melatonin on the reproductive morphology of the hamster might be indirect due to inhibition of gonadotropin secretion (Reiter et al., 2009), melatonin also exerts direct inhibitory effects on the peripheral reproductive organs (Reiter et al., 2013a).

Melatonin might affect the secretory activity of the parathyroid gland. The in vitro melatonin treatment of these glands significantly reduces the cisternae of the RER compared with those of control parathyroid glands (Shoumura et al., 1992). These changes are hypothesized to be induced by melatonin suppressing the synthesis of parathyroid hormone in cells (Shoumura et al., 1992). In human breast cancer cells, specifically MCF‐7 cells, 4 days of exposure to melatonin resulted in the disruption of mitochondrial cristae and vesiculation of the SER (Hill and Blask, 1988). Thus, under normal conditions, the ultrastructure and physiological function of the ER in target organs are changed as a consequence of melatonin treatment.

Melatonin rectifies the abnormal appearance of the ER in target glands

The RER is decreased in volume in the neurons of patients with Alzheimer's disease (AD), and this effect is reversed by melatonin supplementation (Ling et al., 2009). Another study found that unglycosylated ER‐bound β‐APP derivatives are the predominant forms that are marginally affected by melatonin treatment (Lahiri, 1999). Under pathological conditions, melatonin rectifies not only the amount of ER, but also the ultrastructural appearance and presumed function of the ER. Similarly, dilation of the RER is observed in experimental models of acute pancreatitis (Esrefoglu et al., 2006), X‐ray‐irradiated intestine (Hussein et al., 2008) and water avoidance stress‐induced degeneration of the liver parenchyma (Contuk et al., 2005). Dilation of the ER also occurs in the testicular damage associated with hyperlipidaemia (Zhang et al., 2012). Melatonin administration prevents these changes in the ER. Methanol intoxication‐induced liver injury results in the appearance of extensive tubules of the SER, an effect that is also prevented by melatonin administration (Koksal et al., 2012).

Melatonin regulates calcium homeostasis via the ER

An elevation in the intracellular concentration of Ca^{2 +} is considered a common pathological precursor (Santofimia‐Castano et al., 2014), and the ER is the major inositol trisphosphate (IP3)‐sensitive Ca^{2 +} store and plays an important role in the homeostasis and function of Ca^{2 +} (Alves et al., 2011). Cholecystokinin octapeptide (CCK‐8) induces Ca^{2 +} mobilization in mouse, freshly isolated pancreatic acinar cells (del Castillo‐Vaquero et al., 2010). However, melatonin reduces Ca^{2 +} release and modulates the pancreatic responses induced by CCK‐8, which might be explained by the stimulation of Ca^{2 +} transport from cells through the plasma membrane and subsequent Ca^{2 +} reuptake into the ER (del Castillo‐Vaquero et al., 2010; Santofimia‐Castano et al., 2014). Melatonin also exerts a dual inhibitory effect on gonadotropin‐releasing hormone‐induced increases in intracellular Ca^{2 +} and the mobilization of Ca^{2 +} from the ER in the pituitary gland (Vanecek, 1999; Watanabe, 1999). These studies demonstrate that the maintenance of Ca^{2 +} in the ER, including both inhibition of its mobilization and the enhancement of its uptake by the ER, is affected by melatonin.

Melatonin also regulates the Plasmodium falciparum cell cycle via a Ca^{2 +}‐dependent pathway (Alves et al., 2011). The Ca^{2 +} responses to melatonin and the uncaging of IP3 are mutually exclusive in infected red blood cells (Alves et al., 2011). Melatonin promotes the generation of IP3 and opens ER‐localized IP3‐sensitive Ca^{2 +} channels in P. falciparum (Alves et al., 2011). These data not only support the assertion that melatonin is involved in Ca^{2 +} signalling but also provide clues regarding the mechanism underlying this effect. In addition, melatonin inhibits intracellular Ca^{2 +} overload‐induced apoptosis. Treatments with a specific inhibitor of cytosolic Ca^{2 +} reuptake, thapsigargin and/or a Ca^{2 +}‐mobilizing agonist, N‐formyl‐methionyl‐leucyl‐phenylalanine, induce mitochondrial membrane depolarization, caspase activation, phosphatidylserine externalization and DNA fragmentation in leukocytes of both young and elderly volunteers; these effects, however, were far more evident in aged leukocytes (Espino et al., 2011). Notably, melatonin treatment substantially preserves the mitochondrial membrane potential, reverses caspase activation, reduces phosphatidylserine exposure and stops DNA fragmentation in leukocytes of both age groups, and these findings suggest that melatonin can delay Ca^{2 +} overload‐induced apoptosis in aged leukocytes, particularly in those of older individuals (Espino et al., 2011).

Melatonin and ER stress

By controlling ER stress, melatonin protects against multiple disorders, including DM, liver diseases, neurological disorders, reproductive system diseases, lung diseases and chemical poisoning (Espino et al., 2011; Zha et al., 2012; Zaouali et al., 2013; Carloni et al., 2014; Romero et al., 2014; Ali and Kim, 2015; Jeong and Park, 2015; Thakor et al., 2015) (Figure 2). Interestingly, melatonin can cooperate with ER stress to promote the apoptosis of cancer cells and inhibit ER stress to attenuate chemotherapy‐associated side effects and chemoresistance.

Figure 2.

Protective effects of melatonin mediated by the regulation of ER stress. Through the regulation of ER stress, melatonin protects against DM, liver diseases, neurological disorders, reproductive system diseases, cancers, lung diseases and chemical poisoning. In most situations, melatonin exerts its protective effects by inhibiting ER stress. However, melatonin can cooperate with ER stress to promote the apoptosis of HCC.

Diabetes mellitus

ER stress has been demonstrated to cause beta cell dysfunction and death (Back and Kaufman, 2012). In rat insulinoma INS‐1E cells, the expression of insulin receptor substrate (IRS) protein is reduced under thapsigargin‐induced ER stress conditions (Yoo, 2013). Melatonin increases IRS protein expression and thereby elevates insulin secretion in a dose‐dependent manner. Melatonin is hypothesized to mediate insulin synthesis during ER stress and to reverse the results of ER stress by activating the extracellular secretion of insulin (Yoo, 2013).

Furthermore, ER stress may be a central feature of peripheral insulin resistance and DM (Ozcan et al., 2004). One of the main symptoms of DM, hyperglycaemia, is known to induce ER stress (Magierowski et al., 2013), and tunicamycin, a chemical trigger for ER stress, reduces insulin‐mediated glucose transport, supporting the role of ER stress in DM. However, melatonin pretreatment inhibits the decrease in insulin‐mediated glucose transport caused by tunicamycin (Quan et al., 2015). ER stress has been linked to insulin resistance in skeletal muscle. Tunicamycin promotes the phosphorylation of PERK in C2C12 cells and activates gene markers of ER stress, including BIP expression and the splicing of XBP1, in a time‐dependent manner (Quan et al., 2015). However, melatonin pretreatment reverses the elevation in PERK phosphorylation, and the activation of BIP expression and XBP1 splicing. Thus, melatonin inhibits the stimulatory effect of tunicamycin in ER stress and insulin resistance in skeletal muscle cells.

Liver diseases

There is plenty of evidence supporting the notion that ER stress contributes to liver diseases, including hepatic steatosis and liver fibrosis (Dara et al., 2011; Jo et al., 2013; Xiong et al., 2014; Koo et al., 2016). Challenge with tunicamycin has been observed to increase the hepatic triglyceride and intracellular calcium levels through the activation of ER stress (Kim et al., 2015). However, melatonin partially disrupts these phenomena and ameliorates ER stress‐mediated hepatic steatosis (Kim et al., 2015), further limiting non‐alcoholic fatty liver disease and its progression to irreversible complications in ob/ob mice (Stacchiotti et al., 2016).

The treatment of animals with carbon tetrachloride results in hepatic fibrosis, as evidenced by the staining of α‐smooth muscle actin (α‐SMA)‐positive cells. Moreover, increases in the expression of the ER stress chaperones CHOP, BIP and GRP94, in the mRNA levels of PERK, ATF6, ATF4, IRE1 and spliced XBP1, and in phospho‐IRE1, ATF6 and phospho‐PERK protein concentrations have been observed. Through immunohistochemical staining of α‐SMA, San‐Miguel et al. (2015) observed that melatonin significantly inhibits the UPR and ER stress and abolishes the increase in hepatic stellate cells.

Hepatocyte apoptosis is a major contributor to hepatic failure (Tunon et al., 2013). Infection with rabbit haemorrhagic disease virus (RHDV) induces fulminant hepatic failure by increasing the expression of CHOP, BIP and GRP94 and the mRNA levels of ATF6, ATF4, IRE1 and XBP1s (Tunon et al., 2013; San‐Miguel et al., 2014). Melatonin ameliorates RHDV‐induced apoptotic liver damage by attenuating ER stress and modulating the three arms of UPR signalling (Tunon et al., 2013; San‐Miguel et al., 2014). Collectively, melatonin exerts protective effects against liver diseases through the inhibition of ER stress.

Neurological disorders

Recent studies have shown the mechanisms of ER stress‐induced neuronal death (Colla et al., 2012; Mercado et al., 2013; Omura et al., 2013). Furthermore, the decreased levels of melatonin in AD patients suggest a potential relationship between melatonin and AD (Zhou et al., 2003; Rosales‐Corral et al., 2012). Concomitant with the decreased serum melatonin, rats exhibit spatial memory deficits, tau hyperphosphorylation at multiple sites and increased expression of ER stress‐related proteins, including BIP and CHOP (Ling et al., 2009). Simultaneous melatonin supplementation results in partial arrest of these molecular impairments, reduced expression of ER stress‐related proteins and inhibition of AD‐associated behaviour.

In dorsal root ganglion explants, arsenite causes neurotoxicity by elevating the levels of ATF6, ATF4, XBP1 and BIP. Melatonin supplementation suppresses arsenite‐induced ER stress and inhibits arsenite‐induced apoptosis, ultimately protecting against neurotoxicity (Lin et al., 2007; Lin et al., 2009). Furthermore, melatonin protects against ER stress and apoptosis induced by methamphetamine in the SH‐SY5Y neuroblastoma cell line (Wongprayoon and Govitrapong, 2016). In neonatal rats subjected to hypoxia/ischaemia (H/I), melatonin administration significantly reduces brain damage (Carloni et al., 2014). The UPR is strongly activated after H/I, and melatonin significantly reduces the neuronal splicing of XBP1 mRNA, the phosphorylation of eIF2α and the expression of the chaperone proteins BIP and Hsp70 observed in the brain after H/I (Carloni et al., 2014). In another study it was demonstrated that melatonin administration effectively reduces maternal LPS‐induced neonatal inflammation and related brain injury through inhibition of ER stress (Carloni et al., 2016). These findings demonstrate that the attenuation of ER stress is involved in the neuroprotective effect of melatonin against neurological disorders.

Reproductive system diseases

Maternal malnutrition has been shown to impair ovarian function, and also restricts fetal growth causing low birth weight and results in an offspring ovarian phenotype characteristic of premature ovarian aging with a reduced ovarian reserve. Thus, Chan et al. (2015) found that the reduction of adult ovarian follicles induced by early life malnutrition might be mediated by increased ovarian ER stress, resulting in elevated follicular apoptosis accompanied by reduced melatonin levels. These changes are associated with a loss of ovarian vessel density and are consistent with an accelerated ovarian aging phenotype. Thus, a decrease in melatonin might play a role in ovarian aging via activation of ER stress. However, possible protection of this process by melatonin supplementation requires further support.

The administration of LPS during pregnancy retards intrauterine growth and induces fetal death. The placenta of pregnant mice injected with LPS displays apparent ER stress, as determined by decreased BIP expression, obvious eIF2α and JNK phosphorylation and increased CHOP expression (Wang et al., 2011). Melatonin significantly alleviates LPS‐induced placental ER stress, ultimately protecting fetuses from LPS‐induced intrauterine growth restriction and fetal death (Wang et al., 2011).

Cadmium (Cd), a testicular toxicant, induces germ cell apoptosis by increasing the spliced forms of XBP1 and BIP and elevating testicular eIF2α and JNK phosphorylation. These results indicate that ER stress and the UPR pathway are activated by Cd. Melatonin almost completely inhibits the ER stress and UPR induced by Cd in the testes and protects germ cells from apoptosis (Ji et al., 2012).

Cancers

ER stress plays an important role in the development of cancer and apoptosis of cancer cells (Zha et al., 2012; Maurel et al., 2014; Wang and Kaufman, 2014). In rats with diethylnitrosamine‐induced hepatocarcinogenesis, melatonin treatment significantly increases the expression of ATF6, CHOP and BIP, which might further promote the incidence of apoptosis (Moreira et al., 2015). Other studies have shown that melatonin can cooperate with inducers of ER stress to promote cancer cell apoptosis. Significant differences in the apoptosis rate are found between HepG2 cells and HL‐7702 cells (normal human hepatocyte cells) after tunicamycin treatment. The apoptosis rate is significantly higher in HepG2 cells, and this elevated rate is accompanied by the up‐regulation of CHOP and a reduction in the Bcl‐2/Bax ratio. Co‐treatment with tunicamycin and melatonin significantly increases the apoptosis rate by elevating the levels of CHOP and reducing the Bcl‐2/Bax ratio, indicating a pro‐apoptotic effect of melatonin in hepatocellular carcinoma (HCC) through cooperation with ER stress (Zha et al., 2012). Furthermore, a pro‐apoptotic action of melatonin has been observed in other cancer cells (Bizzarri et al., 2013). Co‐treatment with melatonin and tunicamycin (induces ER stress) significantly suppresses the survival of B16F10 melanoma cells compared with treatment with melatonin alone (Kim et al., 2014). Further investigations are required to determine whether this cooperation between melatonin and ER stress exists in other tumours.

The chemoresistance of HCC, which has been widely observed and might be associated with multiple cellular responses to environmental stresses, is also reversed by melatonin (Okuyama et al., 2015). Tunicamycin pretreatment of HepG2 and SMMC‐7721 cells (two human HCC cell lines) markedly decreases the apoptosis rate induced by doxorubicin, indicating a negative role of ER stress in drug resistance. Interestingly, pretreatment with a combination of tunicamycin and melatonin significantly increases the apoptosis induced by doxorubicin, and this finding supports the hypothesis that melatonin attenuates ER stress‐induced resistance to doxorubicin in human HCC cells (Fan et al., 2013).

Cyclosporine A (CsA) is a powerful immunosuppressive drug that has a variety of side effects, including the induction of ER stress, which can further induce autophagy. In the presence of CsA, the expression of catalase is decreased compared with that found in untreated cells, and the levels of BIP and IRE1α are elevated compared with the corresponding levels in untreated cells. Co‐treatment with melatonin inhibits BIP and IRE1α expression, thereby contributing to the alleviation of CsA‐associated side effects (Yoo and Jeung, 2010). Collectively, the results show that, on the one hand, the cooperation between ER stress and melatonin promotes the apoptosis of cancer cells and that, on the other hand, the inhibition of ER stress by melatonin significantly attenuates chemoresistance and the immunosuppressive response.

Lung diseases

The chemotherapeutic agent bleomycin induces lung fibrosis, and melatonin significantly attenuates the bleomycin‐mediated epithelial–mesenchymal transition to myofibroblasts, as evidenced by its repression of α‐SMA expression (Zhao et al., 2014; Yu et al., 2015). Furthermore, melatonin markedly attenuates bleomycin‐induced BIP up‐regulation, elevation of cleaved ATF6 in the lungs and activation of pulmonary eIF2α. These findings indicate the role of ER stress in the protection against lung fibrosis induced by melatonin.

Chemical poisoning

Exposure to the herbicide atrazine adversely affects animal and human health, particularly due to its immunotoxicity. Excessive ER stress is triggered by atrazine through ATF6α, spliced XBP1 and CHOP overexpression in murine splenocytes, but this excessive ER stress is reversed by melatonin (Sharma et al., 2014). Melatonin has also been demonstrated to protect against Cd‐induced testicular toxicity (Ji et al., 2012) and arsenite‐induced neurotoxicity (Lin et al., 2007; Lin et al., 2009) through inhibition of ER stress and the UPR. Furthermore, the possible mechanisms underlying the protective effects of melatonin might also involve modification of the hepatic ER and changes in the metabolism of xenobiotic chemicals (Dhami et al., 1997).

The results from these studies provide new insights into the molecular pathways responsible for the protective effect exerted by melatonin in multiple disorders. Melatonin might be useful as a pharmacological agent that protects against various disorders and promotes optional health by regulating ER stress.

Prospects and conclusion

Melatonin is an agent that regulates protein secretion by influencing various organelles. As a functional organelle, the ER is only one of the targets of melatonin, which also include the mitochondria and Golgi complex (Redondo et al., 2010; Koksal et al., 2012; Zhang et al., 2012). Melatonin treatment could play an important role in improving the histophysiological function of the pineal gland, including the mitochondria, RER and Golgi complex, as observed in goat kids subjected to stress due to premature weaning (Redondo et al., 2010). In addition to extensive SER tubules, increased mitochondria, elevated primary lysosomes and some marked openings of the bile canaliculi are also observed during methanol intoxication, and these effects can be prevented by melatonin (Koksal et al., 2012). In the testicular damage associated with hyperlipidaemia, vacuolar degeneration of the mitochondria and dilation of the ER are observed, and the numbers of mitochondria and lipid droplets are decreased significantly in Leydig and Sertoli cells; these effects were also reversed by melatonin (Zhang et al., 2012). However, further investigations are required to determine whether melatonin specifically influences the ER.

The ER not only participates in the secretion of melatonin but can also be regulated by melatonin in the pineal gland and other tissues, including the reproductive gland, parathyroid gland, pancreas, pituitary, breast cancer cells and leukocytes. Thus, melatonin increases its activation in the pineal gland but reduces this process in other targets. The abnormal ultrastructure and volume of the ER resulting from stress can be reversed by melatonin. Furthermore, melatonin maintains calcium homeostasis via the ER (Figure 3). Thus, in addition to being a well‐known participant of melatonin production and secretion, the ER is also the intracellular regulatory target of melatonin, but the underlying mechanisms require further investigation. In particular, although the protective actions of melatonin are either mediated by its receptors or receptor‐independent (Cardinali et al., 1997; Reiter et al., 2007; Miettunen and Raevuori, 2012), whether the regulation of ER and ER stress by melatonin is mediated by its receptors still needs further investigation through pharmacological methods (Alexander et al., 2015a).

Figure 3.

Involvement of melatonin in ER homeostasis. Melatonin is secreted by the ER and maintains the morphology and function of the ER in the pineal gland. Under stress conditions, including AD, acute pancreatitis, X‐ray‐irradiated intestines, water avoidance‐induced degeneration of the liver parenchyma and hyperlipidaemia‐induced testicular damage (indicated as 1 in the figure), the ER in the pineal gland is dilated and has a decreased volume, and these effects are rectified and increased by melatonin. In target glands and cells, melatonin inhibits the function of the ER and rectifies the decreased number and dilation of the RER and the dilation of tubules in the SER induced by stress conditions, including liver injury induced by methanol intoxication (indicated as 2 in the figure). Furthermore, melatonin inhibits the mobilization of Ca^{2 +} from the ER but promotes the reuptake of Ca^{2 +} into the ER.

After understanding the protective effects of melatonin, it is important to focus on its regulation. Huang et al. (2010) found that 12‐h‐light:12‐h‐dark (LD) and continuous‐light (LL) regimes have opposite effects on the plasma level of melatonin. LL regimes are characterized by increased pineal clock and decreased liver clock genes compared with LD regimes. Under LL regimes, the clock and nuclear transcription factor mRNAs, but not pineal ER stress‐related genes, do not show any daily variations in the organs studied (Huang et al., 2010). Thus, the daily routine might play an important role in maintaining the plasma level of melatonin, normal levels of expression of clock and incidence of ER stress. Regulating the levels of melatonin and ER stress by controlling the day length might serve as a basic contributor to health. Sleep deprivation is one reason for decreased melatonin, and the systemic administration of melatonin appears to constitute a potential neuroprotective treatment against neuronal damage induced by sleep deprivation (Lan et al., 2001). The potential treatment options for restoring a normal sleep cycle include the use of melatonin or its analogues (Reiter et al., 1983; Reiter et al., 2013b) and cognitive behavioural therapy. In modern life, cognitive behavioural therapy might be useful for increasing the production of melatonin and ultimately for regulating ER stress and protecting against various disorders.

Melatonin is a type of indolamine that was initially found to be produced in the pineal gland and is now known to be also synthesized in a variety of other tissues. Rigorous investigations have found that the amount of melatonin produced by enterochromaffin‐like cells in the gastrointestinal tract is approximately 400‐fold greater than that normally produced by pinealocytes (Magierowski et al., 2013; Bertrand et al., 2014). Whether melatonin from tissues other than the pineal gland can be stimulated by any means will be determined by further research. As an endogenous compound, melatonin is safe and presents little evidence of toxicity, but additional investigations are required to determine whether it interacts with other transmitters or hormones.

Author contributions

D.W. and Y.Y. designed the manuscript. W.H., C.F. and S.J. collected the data for the review. W.H., S.D. and L.Y. wrote the paper. Z.M. and W.H. illustrated the manuscript.

Conflict of interest

The authors declare no conflicts of interest.

Hu, W. , Ma, Z. , Di, S. , Jiang, S. , Li, Y. , Fan, C. , Yang, Y. , and Wang, D. (2016) Snapshot: implications for melatonin in endoplasmic reticulum homeostasis. British Journal of Pharmacology, 173: 3431–3442. doi: 10.1111/bph.13651.