Plasmodium spp., which are the causative agents of malaria, are obligate intracellular protozoan parasites. Anand and Babu (2013) reported that experimental cerebral malaria (ECM), caused by P. berghei ANKA (PbA) infection, was related to ER stress. They found that PbA infection-induced ER stress could cause the apoptosis of neuronal cells in mice by activating the three branches of UPR—PERK-eIF2α-ATF4/GADD34, IRE1-XBP1s, and ATF6—along with upregulating the levels of CHOP, cleaved caspase-3 and caspase-12 and downregulating the expression of Bip, calreticulin, and calnexin.

Trypanosome cruzi is the causative pathogen of Chagas disease in humans. Reportedly, the trypomastigotes of T. cruzi infection could induce ER stress in the heart of mice, with an increase in the levels of Bip, PERK, eIF2α, ATF4, and CHOP, thereby causing damage to the host. Interestingly, 2-aminopurine (2-APB, an ER stress inhibitor) treatment could alleviate the pathological damage to the heart by decreasing the phosphorylation of eIF2α and its downstream signaling. Therefore, this indicates that inhibition of ER stress may be a therapeutic target for cardiomyopathy in Chagas patients (Ayyappan et al., 2019).

Toxoplasma is an obligate intracellular parasite and opportunistic pathogenic parasite (Sullivan et al., 2004). Toxoplasma encephalitis is the most serious outcome of toxoplasmosis, which may be fatal to immunocompromised individuals. Some studies have found that Toxoplasma encephalitis was related to ER stress. It has been reported that the tachyzoites of T. gondii RH strain and TgCtwh3 (a representative Chinese 1 Toxoplasma strain) induced apoptosis of neural stem cells and neural stem cell line C17.2 by activating CHOP, caspase-12, and JNK (Wang et al., 2014; Zhou et al., 2015). Pretreatment with tauroursodeoxycholic acid (TUDCA, an ER stress inhibitor) and Z-ATAD-FMK (a caspase-12 inhibitor) led to the inhibition of apoptosis (Wang et al., 2014; Zhou et al., 2015), which suggested that neural stem cell apoptosis induced by both TgCtwh3 and RH strain infection was dependent on the ER stress pathway, and ER stress inhibitors could be used to alleviate Toxoplasma encephalitis. In addition, Wan et al. (2015) showed that virulence factor rhoptry protein 18 (ROP18) secreted by T. gondii was involved in nerve cell apoptosis via the ER stress pathway, characterized by an increase in the expression of cleaved caspase-12, CHOP, and cleaved caspase-3. Ran et al. further indicated that ROP18 induced apoptosis of neural cells by phosphorylating reticulon 1-C [RTN1-C, a protein localized in the ER that is preferentially expressed in the neural cells of the central nervous system (CNS) at Ser7/134 and Thr4/8/118], which led to the acetylation of GRP78 and induced ER stress (An et al., 2018). These results suggest that inhibition of ROP18 of T. gondii can be used as a drug target for the treatment of Toxoplasma encephalitis to inhibit the ER stress-induced apoptosis of host cells.

Schistosoma japonicum is the causative agent of schistosomiasis. The pathogenic mechanism of schistosomiasis is primarily attributed to egg-induced hepatic granuloma and fibrosis and cirrhosis (Yu et al., 2016; Duan et al., 2019). Duan et al. (2019) showed that the level of CHOP, a vital factor in the ER stress-mediated apoptosis pathway, was significantly increased in mice at 6 and 10 weeks following infection with S. japonicum. The study indicated that ER stress may be involved in S. japonicum infection-induced hepatic fibrosis. Moreover, Yu et al. (2016) showed that treatment with taurine, an inhibitor of ER stress, significantly suppressed the egg-induced hepatic granuloma and alleviated hepatic fibrosis in mice at 8 weeks post-infection, along with marked reduction of the expression of GRP78. Therefore, ER stress inhibitors may be a therapeutic drug for hepatic fibrosis.

The summary of ER stress in hosts caused by parasitic infection is shown in Figure 2. Therefore, the UPR signaling pathway may be a therapeutic target to alleviate pathological symptoms.

Plasmodium has a complicated life cycle, including the merozoite, ring, trophozoite, schizont, and gametophyte stages in humans and the ookinete and sporozoite stages in mosquitoes. Chaubey et al. (2014) showed that Plasmodium falciparum lacked the orthologs of XBP1, IRE1, ATF6, and ATF4, and only retained the PERK-eIF2α pathway to regulate translation under ER stress. Three eIF2α kinases have been identified, namely IK1, IK2, and PK4 (eIF2α kinase of Plasmodium (Möhrle et al., 1997), a PERK homolog of mammals) (Ward et al., 2004). It has been reported that increased phosphorylation of eIF2α leads to reduced levels of protein translation, which is associated with the formation of P. falciparum gametophytes and the conversion of the P. berghei gametophytes into ookinetes when treated with dithiothreitol (DTT) (Chaubey et al., 2014; Duran-Bedolla et al., 2017). In addition, Zhang et al. (2012) have shown that PK4 was involved in the invasion of new red blood cells of merozoite-containing schizonts and the gametocyte infecting Anopheles mosquitoes. The inhibition of PK4 of P. berghei by generating a PK4 conditional mutant (PbPK4cKO) would alleviate the symptoms of malaria and inhibit disease transmission. Another study indicated that treatment of GSK2606414 (a small molecule inhibitor of PERK (Axten et al., 2012), which specifically inhibits PK4 instead of IK1 and IK2 in vitro) could block the transformation of P. falciparum from trophozoites to schizonts (Zhang et al., 2017). The transformation between different forms increased the ability of translational regulation of Plasmodium. In addition, Chen et al. (2018) reported that apoptozole, a novel chemical scaffold, was lethal to the chloroquine-sensitive and chloroquine -resistant P. falciparum parasite strains by inhibiting GRP78 function in vitro. Compared to human GRP78, P. falciparum GRP78 showed a lower affinity to the endogenous ligands, ADP and ATP, which indicated that the competitive inhibitors of GRP78 can be investigated for P. falciparum control (Chen et al., 2018).

According to the above mentioned studies, it appears that the PK4-eIF2α pathway plays an important role in both morphological transformation and host transmission in Plasmodium. Thus, PK4 inhibition would inhibit the development of Plasmodium, which implies that PK4 inhibitors may be a potential target in malaria treatment. However, Bridgford et al. (2018) found that dihydroartemisinin (DHA) increased the toxicity to Plasmodium by prolonging PK4 activation and eIF2α phosphorylation. Therefore, appropriate ER stress is beneficial to the development of Plasmodium, while excessive ER stress would be lethal to the parasites.

Leishmania is the pathogen causing Leishmaniasis and has two forms—promastigote and amastigote. Gosline et al. (2011) proved that Leishmania lacked a transcriptional regulation response to UPR, and only retained the translational regulation in ER stress. They also showed an increased level of phosphorylation of eIF2α in L. donovani after treatment of DTT (Gosline et al., 2011). Moreover, Chow et al. (2011) found that the PERK homolog of Leishmania largely colocalized with Bip in ER, which can phosphorylate eIF2α at threonine 166. They further confirmed that PERK-dependent eIF2α phosphorylation was vital for Leishmania to switch from the promastigote to amastigote form in vitro (Chow et al., 2011). Unlike host macrophages having intact UPR pathway, the mere presence of the PERK pathway in L. donovani promoted the parasite’s susceptibility to DTT-induced ER stress (Gosline et al., 2011), which suggests that inhibition of the PERK pathway and induction of ER stress in Leishmania are both potential targets to kill the parasite. Dolai et al. (2011) proved that tunicamycin treatment induced apoptosis of Leishmania major, with an increase in the level of Bip.

Trypanosome brucei is a protozoan parasite that cycles between the tsetse fly (procyclic form) and mammalian host (blood stream form), which causes African sleeping sickness in humans and nagana in livestock (Zhang et al., 2019). Goldshmidt et al. (2010) reported that the expression of Bip of T. brucei was increased in both procyclic and blood stream forms in DTT-induced ER stress, and irrecoverable ER stress could induce spliced leader RNA silencing pathway (SLS pathway, a unique process in T. brucei), which may accelerate programmed cell death (PCD). Besides, Messias Sandes et al. (2019) showed that both DTT and tunicamycin could induce PCD in T. cruzi.

There were three putative eIF2α kinases (TbeIF2K1-K3) in T. brucei, though its genome lacked the homologs of IRE1/XBP1. It was reported that TbeIF2K2, a transmembrane glycoprotein expressed both in the procyclic and bloodstream forms of Trypanosome (Moraes et al., 2007), shared no similar sequence with known eIF2 kinases of mammals and was localized to the flagellar pocket, where endocytosis and exocytosis occur, and all proteins were transported from the flagellar pocket to the cell membrane (Gull, 2003). Therefore, the localization of TbeIF2K2 indicated that it could sense proteins and regulate protein synthesis near the flagellar pocket of the Trypanosome (Moraes et al., 2007), which suggests that TbeIF2K2 may be a good drug target to destroy T. brucei. In addition, Hope et al. (2014) showed that SEC63 (a factor participating in protein translocation machinery in ER) silence-induced ER stress could activate PK3 (TbeIF2K3) and trigger the release of PK3 from the ER to nucleus in the procyclic form of T. brucei. The deletion of PK3 reduced the death of T. brucei in SEC63 silence-induced ER stress, which suggests that PK3 is required for ER stress-induced PCD. Thus, the results indicate that TbeIF2K2 and TbeIF2K3 could be potential drug targets to eliminate T. brucei.

The PERK-eIF2α pathway is also involved in the form transformation of T. cruzi at different developmental stages. Tonelli et al. (2011) reported that the differentiation of non-infective epimastigotes into infective metacyclic trypomastigotes in T. cruzi requires the phosphorylation of Tc-eIF2α.

In conclusion, the results show that eIF2α phosphorylation plays an important role in the survival and development of Trypanosome, while excessive ER stress induced by DTT or tunicamycin can lead to the death of Trypanosome.

Toxoplasma shows two forms in the human host: tachyzoite (a rapidly growing form) and bradyzoite (a quiescent cyst form) (Black and Boothroyd, 2000; Sullivan et al., 2004). It has been reported that T. gondii lacked the homologs of IRE1 and ATF6 (Joyce et al., 2013), while it possessed four TgIF2α kinases, namely TgIF2K-A, TgIF2K-B, TgIF2K-C, and TgIF2K-D (Narasimhan et al., 2008; Konrad et al., 2014). Narasimhan et al. (2008) showed that only TgIF2K-A was a transmembrane protein localized in the ER and bonded to Bip under unstressed conditions. When ER stress occurred, the binding of Bip to TgIF2K-A was reduced, similar to the binding of BiP to PERK in mammals, which suggests that part of the UPR was conserved in T. gondii (Narasimhan et al., 2008).

ER stress is also involved in the differentiation of Toxoplasma. Narasimhan et al. (2008) reported that the phosphorylation of TgIF2α induced by tunicamycin treatment resulted in the differentiation of T. gondii from tachyzoite to bradyzoite cysts. Treatment with salubrinal, an inhibitor of eIF2α dephosphorylation, could also induce the differentiation of bradyzoite cysts, which indicated that TgIF2α phosphorylation was involved in the differentiation of bradyzoite cysts (Narasimhan et al., 2008). Cyst formation is a good way to escape from the host’s immune attack. Therefore, the formation of bradyzoite cysts induced by TgIF2α phosphorylation promotes the survival of T. gondii under stressful conditions. Similar results were confirmed by Joyce et al. (2013). Besides, Joyce et al. (2010) reported that the TgIF2α mutant strain of Toxoplasma (i.e., TgIF2α-S71A, which cannot be phosphorylated) showed a lower virulence to the host cell, a lower survival rate and a slower transmitting speed, compared with the control strain of Toxoplasma. Moreover, Augusto et al. (2018) showed that specific inhibition of TgIF2K-A with GSK2606414 could inhibit the lytic cycle of tachyzoites, including attachment/invasion, replication, egress, and differentiation, which prolonged the survival time of mice with acute toxoplasmosis at a lethal dose of 100 RH strain tachyzoites. Interestingly, GSK2606414 did not show apparent detrimental effects on the host cell though with a high concentration in vitro. Therefore, the results suggest that TgIF2K-A and TgIF2α can be used as drug targets to inhibit Toxoplasma survival.

However, DTT treatment and stearoyl-coenzyme A (CoA) desaturase (SCD) accumulation at the ER could trigger ER stress with increasing phosphorylation of TgIF2α and mediated the apoptosis or autophagy of T. gondii (Nguyen et al., 2017; Hao et al., 2019). Therefore, although the TgIF2K-A/TgIF2α pathway plays a protective role in T. gondii under stress conditions, severe disruption of ER homeostasis can lead to the death of T. gondii.

Entamoeba histolytica infection, caused by ingestion of cysts in contaminated water and food, usually induces amoebic dysentery and liver abscesses in humans (Pineda and Perdomo, 2017). Santi-Rocca et al. (2012) found that no genes encoded the orthologs of PERK and ATF6 in E. histolytica amoeba, while the expression of gene encoding eIF2α was upregulated upon treatment with nitric oxide (NO). Hendrick et al. (2016) showed that eIF2α could be phosphorylated at serine-59 in E. histolytica, with a decrease in translation levels during long-term serum starvation, long-term heat shock, and oxidative stress instead of short-term serum starvation, short-term heat shock, and glucose deprivation, and the viability of EheIF2α-S59D (a phosphomimetic variant of eIF2α) was significantly increased during long-term serum starvation. This study suggests that EheIF2α phosphorylation promotes the survival of E. histolytica under stress conditions. DTT treatment can also induce distinct fragmentation of ER and phosphorylation of EheIF2α, while treatment with SNP and DPTA-NON-Oate (NO donors) did not induce phosphorylation of EheIF2α (Walters et al., 2019). Besides, Kumari et al. (2018) identified the ortholog of IRE1 in E. histolytica (EhIre1) and reported that treatment with tunicamycin resulted in the upregulation of EhIre1. In addition, the level of eIF2α phosphorylation was increased during encystation of Entamoeba invadens, but whether eIF2α is necessary for encystation still needs further investigation (Hendrick et al., 2016).

Echinococcus granulosus is the causative cestode of hydatidosis or cystic echinococcosis (CE) and is a worldwide zoonotic infection that affects many organs in human and mammals (Loos et al., 2018). Nicolao et al. (2017) have identified the ortholog of IRE2, XBP1, and ATF6 in the genome of E. granulosus, but the ortholog of PERK/ATF4 was not found. Treatment with bortezomib (a proteasome inhibitor) led to lower viability of E. granulosus in the larval stage in vitro than that in the control group, with an increase of EgGRP78 and EgIRE2/EgXBP1 mRNA levels in protoscoleces; however, no changes were found in the metacestodes (Nicolao et al., 2017). Another study also showed that arsenic trioxide (As₂O₃) could disturb the intracellular Ca²⁺ homeostasis and activated ER stress-related apoptosis of protoscoleces in vitro, with an increase in the expression of GRP78, caspase-3, and caspase-12 (Li et al., 2018). These studies show that the induction of ER stress can lead to the apoptosis of protoscoleces in vitro.

In sum, the components of the UPR response such as the PERK-eIF2α pathway of some parasites (Figures 3, 4), including Plasmodium, Leishmania, Trypanosome, Toxoplasma, and E. histolytica, play an important role in their survival and development. However, excessive ER stress could induce the death of parasites such as Plasmodium, Leishmania, Trypanosome, Toxoplasma, and E. granulosus (Figure 4). Considering the toxicity of commonly used ER stress inducers such as DTT and tunicamycin, it is difficult to use them to kill parasites in vivo. For those parasites that are more sensitive to ER stress inducers than their hosts, it is necessary to explore the appropriate concentration of these inducers. TUDCA, a bile salt and chemical chaperone used to treat biliary cirrhosis clinically (Lazaridis et al., 2001), partially inhibits ER stress by lowering the levels of PERK, Bip (Malo et al., 2010; Liu et al., 2015; Li et al., 2019). Thus, TUDCA may be an alternative therapy for parasitosis.

Endoplasmic reticulum-associated degradation is another way for maintaining ER homeostasis, which can degrade misfolded protein (Hwang and Qi, 2018). And ERAD also exists in parasites, such as trypanosomes (Tiengwe et al., 2016). In addition, some apicomplexan parasites, including P. falciparum, T. gondii and cryptosporidium, harbor an apicoplast, which is important for parasite survival (Agrawal et al., 2013). Reportedly, ERAD components were associated with importing the apicoplast protein, and lose of ERAD components would lead to the death of parasites (Agrawal et al., 2009; Spork et al., 2009). Thus, apicoplast is a potential anti-parasitic drug target. Ubiquitin-dependent ERAD is essential for the survival of Plasmodium (Chung et al., 2012). Harbut et al. found that the inhibitors of signal peptide peptidase (SPP, a protein of ERAD) was lethal to P. falciparum (Harbut et al., 2012). It was reported that NITD731, a SPP inhibitor, was effective against T. cruzi and T. gondii, and it showed no toxicity to human cell lines (Harbut et al., 2012). Above studies further showed that parasites were much more sensitive to the disruption of protein homeostasis. Thus, inhibition of the two key quality-control mechanisms, UPR and ERAD, may be a potential way for parasites control.