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Cells 2012, 1(4), 738-753; doi:10.3390/cells1040738

Review
Intricately Regulated: A Cellular Toolbox for Fine-Tuning XBP1 Expression and Activity
Andrew E. Byrd and Joseph W. Brewer *
Department of Microbiology and Immunology, College of Medicine, University of South Alabama, Mobile, AL 36688, USA; Email: eab303@jaguar1.usouthal.edu
*
Author to whom correspondence should be addressed; Email: jbrewer@southalabama.edu; Tel.: +1-251-414-8086; Fax: +1-251-460-7931.
Received: 10 August 2012 / Accepted: 25 September 2012 / Published: 15 October 2012

Abstract

: Stress in the endoplasmic reticulum (ER) triggers the unfolded protein response (UPR), a signaling mechanism that allows cellular adaptation to ER stress by engaging pro-adaptive transcription factors and alleviating protein folding demand. One such transcription factor, X-box binding protein (XBP1), originates from the inositol-requiring transmembrane kinase/endoribonuclease 1 (IRE1) UPR stress sensor. XBP1 up-regulates a pool of genes involved in ER protein translocation, protein folding, vesicular trafficking and ER- associated protein degradation. Recent data suggest that the regulation of XBP1 expression and transcriptional activity may be a tissue- and stress-dependent phenomenon. Moreover, the intricacies involved in “fine-tuning” XBP1 activity in various settings are now coming to light. Here, we provide an overview of recent developments in understanding the regulatory mechanisms underlying XBP1 expression and activity and discuss the significance of these new insights.
Keywords:
endoplasmic reticulum; endoplasmic reticulum stress; unfolded protein response; XBP1

1. Introduction

Cells are highly sensitive to conditions that disrupt the environment of the endoplasmic reticulum (ER) or that increase demand on its machinery for synthesis, maturation and transport of secretory cargo. Various physiological and pathophysiological conditions interfere with protein folding processes in the ER, leading to accumulation of misfolded or unfolded proteins, a cellular condition referred to as “ER stress”. Adapting to the nature, intensity, and duration of ER stress, cells launch the unfolded protein response (UPR), a tri-partite intracellular signaling mechanism that triggers translational control and an extensive transcriptional response to balance client protein load with the folding capacity of the ER [1]. The three distinct mammalian UPR signaling pathways are initiated by the ER transmembrane sensors protein kinase RNA-activated (PKR)-like ER kinase (PERK), activating transcription factor 6 (ATF6), and inositol-requiring enzyme 1 (IRE1). Activated PERK phosphorylates the α subunit of eukaryotic initiation factor 2 (eIF2α), effectively down-regulating protein synthesis [2,3,4,5]. Proteolytic processing of ATF6 yields an active transcription factor (ATF6α(N)) that up-regulates expression of ER resident quality control proteins, including chaperones and ER-associated degradation (ERAD) components [6,7,8,9,10]. Upon activation of IRE1, its endoribonucleolytic activity catalyzes an unconventional cytosolic splicing of X-box binding protein 1 (XBP1) mRNA, resulting in a translational frameshift that generates XBP1S, a pro-adaptive, basic-region leucine zipper (bZIP) transcription factor [11,12,13]. If these adaptive mechanisms do not sufficiently recover ER homeostasis, the UPR can commit the damaged cell to death by apoptosis [14].

XBP1 is subject to transcriptional, post-transcriptional and post-translational control mechanisms [15,16,17,18,19,20,21,22,23,24,25,26], indicating that the activity of this crucial UPR transcription factor is carefully balanced. Indeed, XBP1 has tissue-distinct roles during the UPR in different physiological processes and diseases. XBP1, a member of the CREB/ATF (cyclic AMP response element binding protein/activating transcription factor), family of transcription factors, was originally identified as a protein that binds to the HLA (human leukocyte antigen) DRα promoter [27]. Since then, studies have unveiled multiple roles for XBP1 in diverse processes including secretory function, the inflammatory response, lipid metabolism and glucose homeostasis [28]. XBP1 deficiency impairs normal development and function of antibody-producing plasma cells [29] as well as pancreatic, liver and salivary gland cells [30,31,32,33]. XBP1 also plays a role in the development and survival of dendritic cells [34] and in the response of macrophages to certain Toll-like receptor agonists [35]. Defects in the XBP1 gene and/or protein have also been linked to critically important problems in human health such as ulcerative colitis/Crohn’s disease [36], insulin resistance and type 2 diabetes [37].

On the whole, these observations highlight the critical role of XBP1 as a pro-adaptive transcription factor in various physiological processes. Though genetically modified mice have allowed key discoveries of XBP1 function, a better understanding is needed concerning the mechanisms employed by cells to specifically moderate the activity of XBP1.

2. XBP1 Biosynthesis and Function

IRE1 is a bifunctional, single-pass ER transmembrane Ser/Thr kinase and endoribonuclease. This UPR transducer was originally identified in yeast [38,39,40] and is very similar across eukaryotic species. Mammals express two isoforms of IRE1, α and β. IRE1α is ubiquitously expressed [41], whereas IRE1β is limited to the gut epithelium [42]. Here, our discussion will focus on IRE1α since it is expressed in all cell types. In cells experiencing ER stress, active IRE1α catalyzes the cytosolic removal of a 26 nucleotide (nt) intron from XBP1 pre-mRNA (XBP1u) (Figure 1b). This unconventional splicing event results in a translational frameshift that joins the two open reading frames (ORFs) encoding a DNA binding domain (DBD) and a transcriptional activation domain (AD) [11,12,13] (Figure 1a). Translation of spliced XBP1 mRNA (XBP1s) generates XBP1S, a bZIP transcription factor that binds to and activates two cis-acting promoter motifs, the ER stress response element (ERSE) and unfolded protein response element (UPRE) [13]. In addition to activating its own transcription, XBP1S induces genes encoding: factors involved in translocation of nascent polypeptides into the ER (SRP54, SEC61A, SEC61G and TRAM1); ER chaperones (ERdj4, ERdj5, HEDJ, GRP58 and PDI-P5), ERAD components (EDEM, OS9, HERP, and p58IPK); factors involved in vesicular transport (SEC23B and SEC24C)) [43,44]. More recently, XBP1S was shown to up-regulate expression of a microribonucleic acid (miRNA), miR-346, that targets the human antigen peptide transporter 1 (TAP1) mRNA [45].

Complete deletion of XBP1 in mice results in embryonic lethality at ~13 weeks gestation [33]. Rescue of embryonic lethality by targeting an XBP1 transgene selectively to hepatocytes led to early post-natal lethality via activation of ER stress-mediated proapoptotic pathways [30]. Specifically, the phenotype consisted of weak expression of ER chaperone genes and poorly developed ER in pancreatic and salivary gland acinar cells, and this correlated with impaired production of pancreatic digestive enzymes [30]. Similarly, XBP1S is vital for ER expansion and induction of high-rate immunoglobulin synthesis during plasma cell differentiation [44,46].

3. Post-transcriptional Modulation of XBP1 Expression

Recent reports indicate that post-transcriptional mechanisms influence the fate of XBP1 mRNA. Regulatory mechanisms implicated include unique localization of XBP1 mRNA at the ER membrane and translational pausing that facilitates IRE1α-dependent splicing. In addition, XBP1 mRNA is targeted by miRNA.

IRE1α-mediated splicing of XBP1 mRNA occurs in the cytosol [47,48], in contrast to conventional mRNA splicing that takes place in the nucleus. Only recently have discoveries shed light on underlying mechanisms that orchestrate the localization of XBP1 mRNA within proximity of IRE1α at the ER membrane (Figure 1b). A novel observation of cellular localization of total XBP1 mRNA was reported in a study examining mRNA partitioning and translation in the ER and cytosolic compartments during the UPR [49]. Surprisingly, total XBP1 mRNA was found to be predominantly membrane associated, although its protein products, XBP1U and XBP1S, are soluble [49]. A subsequent study confirmed XBP1u mRNA association with the ER membrane, but reported XBP1s mRNA re-distribution to cytosolic compartments for translation [24]. Yanagitani and colleagues [24] further implicated a conserved hydrophobic region (HR2) near the carboxyl-terminus of XBP1U as an ER membrane association domain (Figure 1a, b). This group speculated that the HR2 of nascent XBP1U polypeptide chains might cotranslationally recruit XBP1u mRNA to the ER membrane as part of a mRNA-ribosome-nascent chain complex (R-RNC) [24] (Figure 1b). In addition, they recently reported that translation of the XBP1u mRNA transiently pauses to stabilize the R-RNC complex [25]. This entire process is dependent on XBP1U sequences that are highly similar across multiple species, specifically the HR2 and an additional region near the carboxyl-terminus [25] (Figure 1a).

While the Stephens [49] and Yanagitani [24,25] studies agree that XBP1u mRNA localizes at the ER membrane, ambiguity remains as to whether XBP1 mRNA shifts from the ER membrane to the cytosol after IRE1α-mediated splicing has occurred. Notably, the two studies were conducted in different cell lines under different strengths of ER stress inducers. Importantly, the HR2 is located within the 3’ segment of the XBP1u coding region where the translational frame is altered by IRE1α−mediated splicing, resulting in XBP1S which lacks the HR2 [24]. Finally, studies of XBP1-deficient mice have revealed hyperactivation of IRE1α associated with splicing of a truncated XBP1 mRNA in liver and intestinal tissue [32,36], indicating that expression of XBP1U is not required for splicing. Perhaps, the sub-cellular distribution of total XBP1 mRNA is determined in a tissue- and/or stress-specific fashion. Further studies are required to delineate a full understanding of these mechanisms and their relevance in vivo.

More recently, we and others have reported miRNA-mediated regulation of XBP1 expression via the 3’ untranslated region (UTR) of XBP1 mRNA [15,50] (Figure 1b). miRNA are a class of endogenous, non-coding, single-stranded RNAs ~22 nts long that typically function as post-transcriptional repressors of gene expression [51]. Although the specific biological functions of miRNA in ER stress and the UPR remain largely unknown, a few ER stress-inducible miRNAs have been identified [15,45,52].

Our group identified a miRNA, miR-30c-2* (since designated miR-30c-2-3p), that targets a single site in the 3′-UTR of XBP1 mRNA (Figure 1b). Over-expressing miR-30c-2* reduced the levels of XBP1 and its target genes in stressed cells, whereas inhibiting miR-30c-2* activity had the opposite effect, boosting XBP1 levels and promoting cell survival [15]. Induction of ER stress by subjecting human and mouse cell lines to treatment with tunicamycin (Tm), an inhibitor of N-linked glycosylation, triggers the UPR and activates IRE1α, thereby generating XBP1S. Interestingly, miR-30c-2* is up-regulated during the Tm-induced UPR, concomitant with XBP1, suggesting that this miRNA might affect XBP1S expression levels as the UPR proceeds [15]. Our study further showed that up-regulation of miR-30c-2* is dependent on another key signaling component of the UPR, the protein kinase PERK. The mechanism involves the transcription factor nuclear factor-κB (NF-κB) and its interaction with a NF-κB enhancer motif upstream of the miR-30c-2* genes [15]. In the UPR, NF-κB is activated downstream of PERK [53,54]. Importantly, a separate study revealed that phosphorylation of eIF2α is required for optimal induction of XBP1S by a mechanism involving both XBP1 mRNA stabilization and translation inhibition [18]. Therefore, emerging evidence indicates that regulatory cross-talk between the IRE1/XBP1 and PERK pathways influences the strength of XBP1S induction.

Another miRNA, miR-214, was recently implicated as a negative regulator of XBP1 expression in hepatocellular carcinomas (HCC) [50] (Figure 1b). In HCC, miR-214 expression was down-regulated in response to pharmacologic ER stress-inducing agents, hypoxia and lipopolysaccharide stimulation, suggesting that miR-214 represses XBP1 expression until the UPR is activated. Down-regulation of miR-214 correlated with NF-κB activation, whereby NF-κB exerted a negative regulatory affect on expression of the miR-199a/214 gene cluster [50]. However, a direct binding site through which NF-κB mediates this effect was not demonstrated, raising the possibility of an indirect regulatory mechanism.

Cells 01 00738 g001 1024
Figure 1. Regulatory elements of XBP1 expression and activity.(a) Figure of XBP1 protein variants (XBP1U and XBP1S). Numbers indicate amino acid positions. Basic and leucine zipper (ZIP) domains are indicated as well as ORF1 and ORF2 in the carboxyl-terminal domain of XBP1U and XBP1S, respectively. Hydrophobic region (HR), translational pausing (TP) and “proteasome-susceptible” degradation (PSD) domains of XBP1U are also highlighted. (b) Model for XBP1 mRNA localization and processing at the ER membrane. The ER-localizing HR domain, TP domain, and microRNA targeting regions (MTR) are highlighted. The 26 nt intron spliced from XBP1u mRNA by IRE1α is depicted in red. (c) Post-translational modifications of XBP1S. Acetylation (Ac) by p300 increases stability and enhances transcriptional activity of XBP1S. SUMOylation (SUMO) by PIAS2 decreases activity of XBP1S. p38 MAPK-mediated phosphorylation (P) enhances XBP1S nuclear localization.

Click here to enlarge figure

Figure 1. Regulatory elements of XBP1 expression and activity.(a) Figure of XBP1 protein variants (XBP1U and XBP1S). Numbers indicate amino acid positions. Basic and leucine zipper (ZIP) domains are indicated as well as ORF1 and ORF2 in the carboxyl-terminal domain of XBP1U and XBP1S, respectively. Hydrophobic region (HR), translational pausing (TP) and “proteasome-susceptible” degradation (PSD) domains of XBP1U are also highlighted. (b) Model for XBP1 mRNA localization and processing at the ER membrane. The ER-localizing HR domain, TP domain, and microRNA targeting regions (MTR) are highlighted. The 26 nt intron spliced from XBP1u mRNA by IRE1α is depicted in red. (c) Post-translational modifications of XBP1S. Acetylation (Ac) by p300 increases stability and enhances transcriptional activity of XBP1S. SUMOylation (SUMO) by PIAS2 decreases activity of XBP1S. p38 MAPK-mediated phosphorylation (P) enhances XBP1S nuclear localization.
Cells 01 00738 g001 1024

Together, these findings break the ice for exploration into potential roles for miRNA-mediated regulation of XBP1. However, the in vivo relevance of these mechanisms remains unexplored. It will be interesting to investigate the degree to which miRNA regulate XBP1 translation and affect gene expression, cell function and cell fate in various physiological and pathophysiological conditions involving XBP1, such as plasma cell differentiation [46], macrophage activation [35] and tumor cell survival [55].

4. Post-translational Modifications of XBP1S

A series of recent discoveries have implicated XBP1S as a target for a variety of post-translational modifications, including phosphorylation, SUMOylation, acetylation and deacetylation (Figure 1c). These modifications affect XBP1S turnover, sub-cellular localization and transcriptional activity.

4.1. Phosphorylation

A recent study revealed a novel phosphorylation of XBP1S and linked this modification to the maintenance of glucose homeostasis in mice [17]. Convincing evidence showed that XBP1S is phosphorylated on Thr48 and Ser61 by the ER stress responsive p38 MAPK (mitogen-activated protein kinase) (Figure 1c) and that this modification enhanced its nuclear translocation by 5-fold [17]. The study further addressed whether p38 MAPK modulates XBP1S activity in vivo. In the ob/ob mouse model of obesity, reduced XBP1S activity in the liver has been associated with increased liver tissue-specific ER stress and defective glucose homeostasis [20]. These observations correlate well with defective XBP1S nuclear translocation in the livers of ob/ob mice [20]. Indeed, inhibition of p38 MAPK blocked entry of XBP1S into the nucleus, whereas restoration of p38 MAPK activity in the livers of ob/ob mice enhanced XBP1S nuclear translocation [17]. These data highlight the significance of XBP1S phosphorylation on Thr48 and Ser61 for its nucleocytoplasmic shuttling and demonstrate the impact of this event on cell function in vivo. Perhaps, this novel mechanism of XBP1S post-translational modification via phosphorylation can be exploited for therapeutic treatment of various pathological states including obesity and type 2 diabetes, inflammatory conditions, and multiple myeloma.

4.2. SUMOylation

The post-translational modification of proteins by small ubiquitin-like modifier (SUMO) has been shown to regulate a variety of cellular processes such as signal transduction, protein degradation, nuclear-cytosolic transport and gene transcription [56]. A seminal study showed that two lysine residues, Lys276 and Lys297, near the carboxyl-terminus of XBP1S are subject to covalent SUMOylation [16]. In addition, XBP1S SUMOylation appears to be highly specific for the SUMO E3 ligase PIAS2 (protein inhibitor of activated STAT2) (Figure 1c). These two SUMOylation events exerted an additive negative effect on XBP1S, reducing its transcriptional activity. Conversely, ablation of SUMOylation significantly enhanced XBP1S transcriptional activity. The study further demonstrated that SUMOylation does not alter nuclear translocation of XBP1S [16]. Thus, the mechanism by which SUMOylation dampens XBP1S activity and the significance of this mechanism in physiologic processes involving XBP1S await further investigation.

4.3. Acetylation and Deacetylation

XBP1S can also be targeted for acetylation and deacetylation mediated by p300 and SIRT1 (sirtuin 1), respectively [22]. As p300 increases acetylation of XBP1S, the stability and transcriptional activity of XBP1S are enhanced (Figure 1c). In contrast, SIRT1 deacetylates XBP1S and inhibits its transcriptional activity (Table 1). Consistent with these data, SIRT1−/− MEFs (mouse embryo fibroblasts) subjected to pharmacologic ER stress exhibited greater resistance to ER stress-induced cell death when compared to wild-type MEFs [22]. Therefore, the extent that acetylation and deacetylation influence XBP1S in physiologic settings warrants further study.

5. Protein-protein Interactions of XBP1U and XBP1S

A number of binding partners have recently been identified for the XBP1 proteins, including transcription factors and other types of regulatory proteins. Such protein-protein interactions have been implicated in regulating the turnover, nucleocytoplasmic shuttling and transcriptional activity of XBP1S (Figure 2).

5.1. XBP1U Negatively Regulates XBP1S

The role(s) of XBP1U have not been easily identified. However, Yoshida and colleagues [26] made a key discovery in showing that heterodimerization of XBP1U and XBP1S negatively regulates XBP1S. XBP1U contains a DBD, lacks an AD and is extremely labile [11,13,19,21,57]. In contrast, while XBP1S contains the same DBD, it also harbors an AD and is more stable [11]. However, both XBP1U and XBP1S contain a nuclear localization sequence (NLS) required for nuclear translocation. In addition to the NLS, XBP1U contains two other subcellular localization determinants: a nuclear export sequence (NES) and a NES attenuator [26]. As XBP1U shuttles between the nucleus and the cytoplasm, this factor can bind XBP1S and promote its localization to the cytoplasm [26]. This mechanism appears to facilitate proteasomal degradation of the XBP1U-XBP1S complex (Figure 2). Supporting this idea is the presence of a proteasome-susceptible “degradation domain” located in amino acids 209-261 of XBP1U [26] and recent evidence that XBP1U interacts with 20S proteasomes via its carboxyl-terminus [19]. Though this model warrants further investigation, particularly in the context of physiologic processes, XBP1U appears to function as a repressor of XBP1S activity as the UPR proceeds. This mechanism might play an especially significant role under conditions in which UPR-mediated splicing of XBP1 mRNA is either weakly activated or is waning due to resolution of ER stress.

5.2. p85 Subunit of PI3K Enhances XBP1S Nucleocytoplasmic Shuttling

More recently, two independent studies were simultaneously reported that identified an interaction between the p85α subunit of phosphatidylinositol 3-kinase (PI3K) and XBP1S in the context of metabolic control in diabetes models [20,23]. While both studies utilized in vitro model systems to validate the p85α-XBP1S interaction, Park and colleagues [20] went further to show that the p85β subunit could also form a protein complex with XBP1S. The p85-XBP1S interaction most notably increased XBP1S nucleocytoplasmic shuttling, resulting in increased nuclear accumulation of XBP1S [20,23] (Figure 2). Whether the reported nuclear accumulation is due strictly to enhanced nuclear transport, or whether p85 alters XBP1S stability needs further investigation. p85α and p85β also heterodimerize with each other to form an insulin-sensitive complex that dissociates when cells are exposed to insulin [20], an inducer of PI3K activity. Insulin stimulation also increases XBP1S nuclear translocation, suggesting a model whereby insulin facilitates the dissociation of the p85α-p85β complex, freeing the subunits to interact with XBP1S and enhance its nucleocytoplasmic shuttling [20] (Figure 2). In addition, p85α-deficient brown preadipocyte cells exhibited a defective Tm-induced UPR, including a 40% reduction in nuclear XBP1S relative to wild-type cells [23]. Winnay and colleagues [23] showed that liver-specific deletion of p85α in mice hindered the UPR, XBP1S nucleocytoplasmic shuttling and the ability of the liver to adequately resolve Tm-induced ER stress. Similar observations were made when mice lacking both p85α and p85β in the liver were subjected to refeeding-induced metabolic overload [20]. In insulin resistant ob/ob mice, refeeding-induced metabolic overload was accompanied by defective XBP1S nuclear translocation and diminished induction of ER chaperone genes in liver cells. Notably, this correlated with a reduced level of p85-XBP1S complexes [20]. In contrast, in livers of wild-type mice, XBP1S interacted with p85 subunits and a normal UPR was detectable [20]. These data suggest that p85-XBP1 complexes form in vivo and that this interaction is crucial for metabolic homeostasis in the liver. Specifically, it seems that an intact insulin-sensitive p85 dissociative response is required for the partnering of p85α and p85β with XBP1S, an interaction that optimizes nuclear translocation of XBP1S and ensures an appropriate UPR in the liver during conditions of metabolic overload.

5.3. XBP1S Directs FOXO1 to Proteasome-Mediated Degradation

XBP1S was also reported to physically interact with the transcription factor Forkhead box O1 (FOXO1) [58] (Figure 2). The FoxO subfamily of transcription factors regulates assorted cellular functions, such as differentiation, metabolism, proliferation and survival [59,60]. FOXO1 is a wide-range regulator of energy homeostasis, including hepatic glucose output, adipocyte and muscle differentiation, and feeding behavior in the brain [59,60]. Importantly, FOXO1 promotes development of hyperglycemia in insulin-resistant conditions. XBP1S appears to bind directly to FOXO1 and facilitate its degradation by the 26S proteasome [58]. In addition, enforced expression of a DNA-binding-defective mutant of XBP1S in the liver tissue of ob/ob mice increased glucose tolerance and reduced blood glucose levels [58]. Prior to this report, the relevance of XBP1 in metabolic homeostasis was generally assumed to involve its classical function of improving ER folding capacity, thereby enhancing insulin sensitivity. Remarkably, this study suggests that XBP1S may act to reduce serum glucose concentration and increase glucose tolerance independently of its role in transactivating ER stress responsive genes [58]. These findings open new avenues of study regarding the XBP1S-FOXO1 complex in metabolic disorders like type 2 diabetes and raise the intriguing possibility that XBP1S might also affect other cellular processes via interaction with as yet undefined regulatory factors.

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Figure 2. Binding partners of XBP1 proteins. The transcriptional activity of XBP1S is modulated through protein-protein interactions. The p85α/β subunits of phosphatidylinositol 3-kinase (PI3K) bind to XBP1S and enhance its nuclear translocation. The active form of activating transcription factor 6 that traffics into the nucleus (ATF6(N)) interacts with XBP1S to exert a combinatorial effect on expression of ER-associated degradation (ERAD) components. Proteasome-mediated degradation of XBP1S is enhanced by direct interactions with both XBP1U and the transcription factor Forkhead box O1 (FOXO1).

Click here to enlarge figure

Figure 2. Binding partners of XBP1 proteins. The transcriptional activity of XBP1S is modulated through protein-protein interactions. The p85α/β subunits of phosphatidylinositol 3-kinase (PI3K) bind to XBP1S and enhance its nuclear translocation. The active form of activating transcription factor 6 that traffics into the nucleus (ATF6(N)) interacts with XBP1S to exert a combinatorial effect on expression of ER-associated degradation (ERAD) components. Proteasome-mediated degradation of XBP1S is enhanced by direct interactions with both XBP1U and the transcription factor Forkhead box O1 (FOXO1).
Cells 01 00738 g002 1024
Table Table 1. Molecular mechanisms modulating XBP1 expression and activity. Overview of reported regulatory mechanisms for the XBP1 transcription factor.

Click here to display table

Table 1. Molecular mechanisms modulating XBP1 expression and activity. Overview of reported regulatory mechanisms for the XBP1 transcription factor.
Cellular ProcessEffectorMechanismEffect on XBP1ModelReference
Post-trancriptionIRE1cytosolic mRNA splicingcytosolic mRNA splicing/religation/ORF frameshift pharmacologic-induced ER stress[11,12,13]
HR2 of nascent XBP1UXBP1u mRNA recruitmentlocalize XBP1 mRNA at ER membrane within close proximity of IRE1 in vitro expression of XBP1[24]
XBP1u COOH-terminustranslational pausingstabilize R-RNC complex at ER membrane for efficient targeting and splicing of XBP1u mRNApharmacologic-induced ER stress[25]
miR-30c-2*inhibits XBP1 fine-tune XBP1 expression as the UPR proceedspharmacologic-induced ER stress[15]
miR-214expression inhibits XBP1 repress XBP1 expression prior to UPR activationLPS- and pharmacologic- induced ER stress[50]
Post-translationp38 MAPKphosphorylationincrease XBP1S stability/enhance XBP1S nucleocytoplasmic shuttlingpharmacologic-induced ER stress; ob/ob mice [17]
PIAS2SUMOylationdecrease transcriptional activity of XBP1Sin vitro expression of XBP1; pharmacologic-induced ER stress[16]
p300acetylationincrease XBP1S stabilitypharmacologic-induced ER stress[22]
SIRT1deacetylationdecrease XBP1S stabilitypharmacologic-induced ER stress[22]
Protein interactionXBP1Uheterodimerizationtarget XBP1U-XBP1S complex for proteasome-mediated degradationtransient over-expression in vitro[26]
PI3K p85 subunitsheterodimerizationenhance XBP1S nucleocytoplasmic shuttlingpharmacologic-induced ER stress; metabolic induced overload in ob/ob mice[20,23]
FOXO1heterodimerizationdirect protein complex to 26S proteasome in vitro expression; glucose homeostasis in ob/ob mice[58]
ATF6α( N)heterodimerizationcombinatorially enhance ERAD gene expressionpharmacologic-induced ER stress[10]

5.4. ATF6α Interacts with XBP1S to Optimize ERAD

Another protein-protein interaction involves the combinatorial effect of XBP1S and the UPR transcription factor, ATF6α(N), in the expression of genes involved in ERAD. The transcriptional induction of ERAD components is a primary response to ER stress and aids in ER homeostasis by facilitating the elimination of terminally misfolded proteins. Both XBP1S and ATF6α(N) activate transcription of several ERAD genes via an as yet unidentified promoter element(s). However, in vitro studies revealed cross-talk between these two UPR branches, whereby ATF6α(N) heterodimerizes with XBP1S [10]. ATF6α cannot bind to the UPRE [13], a promoter element recognized by XBP1S and suspected to control expression of some ERAD genes. Thus, it was proposed that ATF6α-XBP1S heterodimers might regulate ERAD genes via the UPRE [10] (Figure 2). In support of this model, induction of some ERAD genes is dependent on both ATF6α and XBP1S [10]. A more recent study found that XBP1U can specifically bind to ATF6α(N) during the late-stage UPR and facilitate rapid degradation of both proteins [61]. This is consistent with a previous report that XBP1U can dimerize with ATF6α(N) [62], though the affect of the interaction was not established at the time. Both studies reported XBP1U interaction to be specific for ATF6α but not ATF4 [61,62], a transcription factor originating from the PERK branch of the UPR. Though these findings suggest a unique cross-talk between the ATF6α and IRE1/XBP1 branches of the UPR, further studies are needed to establish the relevance of these interactions in vivo during physiological and pathophysiological UPR-inducing events.

6. Concluding Remarks

Whether via transcriptional, post-transcriptional, co-translational, post-translational modifications and/or protein-protein interactions (Table 1), examples of mechanisms that can finely adjust XBP1S activity are increasingly evident. In the complexity of UPR signal transduction, the emergence of an IRE1/XBP1 signaling pathway that is subject to such diverse controls leads one to postulate that we have just begun to understand the breadth and importance of XBP1 regulatory activities. The combinatorial effects of these molecular mechanisms may vary widely depending on the tissue-type undergoing ER stress. For example, in the stressful conditions of the tumor microenvironment, different types of ER stressors and/or varying degrees of the same ER stress can be present, such as hypoxia and nutrient deprivation.

An obvious, albeit challenging, direction for this exciting field is to establish the degree to which XBP1 regulatory mechanisms occur and function in vivo. Moving forward, considerable attention should be given to interpreting how these findings translate to our understanding of XBP1 related pathologies and the therapeutic potential of exploiting these mechanisms in certain disease states such as inflammatory disorders [63] and cancer [64]. Further defining the dynamic interplay of XBP1 and the molecular mechanisms that modulate its activity will provide a deeper understanding of the fundamental roles of the UPR, specifically IRE1/XBP1 signaling, in cell physiology.

Acknowledgements

The authors thank the US National Institutes for grant support (GM061970).

Conflict of Interest

The authors declare no conflict of interest.

References

  1. Walter, P.; Ron, D. The unfolded protein response: from stress pathway to homeostatic regulation. Science 2011, 334, 1081–1086. [Google Scholar] [CrossRef]
  2. Harding, H.P.; Zhang, Y.; Bertolotti, A.; Zeng, H.; Ron, D. Perk is essential for translational regulation and cell survival during the unfolded protein response. Mol. Cell 2000, 5, 897–904. [Google Scholar] [CrossRef]
  3. Harding, H.P.; Zhang, Y.; Ron, D. Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature 1999, 397, 271–274. [Google Scholar] [CrossRef]
  4. Scheuner, D.; Song, B.; McEwen, E.; Liu, C.; Laybutt, R.; Gillespie, P.; Saunders, T.; Bonner-Weir, S.; Kaufman, R.J. Translational control is required for the unfolded protein response and in vivo glucose homeostasis. Mol. Cell 2001, 7, 1165–1176. [Google Scholar] [CrossRef]
  5. Shi, Y.; Vattem, K.M.; Sood, R.; An, J.; Liang, J.; Stramm, L.; Wek, R.C. Identification and characterization of pancreatic eukaryotic initiation factor 2 alpha-subunit kinase, PEK, involved in translational control. Mol. Cell. Biol. 1998, 18, 7499–7509. [Google Scholar]
  6. Adachi, Y.; Yamamoto, K.; Okada, T.; Yoshida, H.; Harada, A.; Mori, K. ATF6 is a transcription factor specializing in the regulation of quality control proteins in the endoplasmic reticulum. Cell Struct. Funct. 2008, 33, 75–89. [Google Scholar] [CrossRef]
  7. Haze, K.; Okada, T.; Yoshida, H.; Yanagi, H.; Yura, T.; Negishi, M.; Mori, K. Identification of the g13 (camp-response-element-binding protein-related protein) gene product related to activating transcription factor 6 as a transcriptional activator of the mammalian unfolded protein response. Biochem. J. 2001, 355, 19–28. [Google Scholar] [CrossRef]
  8. Haze, K.; Yoshida, H.; Yanagi, H.; Yura, T.; Mori, K. Mammalian transcription factor ATF6 is synthesized as a transmembrane protein and activated by proteolysis in response to endoplasmic reticulum stress. Mol. Biol. Cell 1999, 10, 3787–3799. [Google Scholar]
  9. Wu, J.; Rutkowski, D.T.; Dubois, M.; Swathirajan, J.; Saunders, T.; Wang, J.; Song, B.; Yau, G.D.; Kaufman, R.J. ATF6α optimizes long-term endoplasmic reticulum function to protect cells from chronic stress. Dev. Cell 2007, 13, 351–364. [Google Scholar] [CrossRef]
  10. Yamamoto, K.; Sato, T.; Matsui, T.; Sato, M.; Okada, T.; Yoshida, H.; Harada, A.; Mori, K. Transcriptional induction of mammalian ER quality control proteins is mediated by single or combined action of ATF6α and XBP1. Dev. Cell 2007, 13, 365–376. [Google Scholar] [CrossRef]
  11. Calfon, M.; Zeng, H.; Urano, F.; Till, J.H.; Hubbard, S.R.; Harding, H.P.; Clark, S.G.; Ron, D. IRE1 couples endoplasmic reticulum load to secretory capacity by processing the XBP-1 mRNA. Nature. 2002, 415, 92–96. [Google Scholar] [CrossRef]
  12. Shen, X.; Ellis, R.; Lee, K.; Liu, C.-Y.; Yang, K.; Solomon, A.; Yoshida, H.; Morimoto, R.; Kurnit, D.M.; Mori, K.; et al. Complementary signaling pathways regulate the unfolded protein response and are required for C. elegans development. Cell 2001, 107, 893–903. [Google Scholar] [CrossRef]
  13. Yoshida, H.; Matsui, T.; Yamamoto, A.; Okada, T.; Mori, K. XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell 2001, 107, 881–891. [Google Scholar] [CrossRef]
  14. Tabas, I.; Ron, D. Integrating the mechanisms of apoptosis induced by endoplasmic reticulum stress. Nat. Cell Biol. 2011, 13, 184–190. [Google Scholar] [CrossRef]
  15. Byrd, A.E.; Aragon, I.V.; Brewer, J.W. MicroRNA-30c-2* limits expression of proadaptive factor XBP1 in the unfolded protein response. J. Cell Biol. 2012, 196, 689–698. [Google Scholar] [CrossRef]
  16. Chen, H.; Qi, L. SUMO modification regulates the transcriptional activity of XBP1. Biochem. J. 2011, 429, 95–102. [Google Scholar] [CrossRef]
  17. Lee, J.; Sun, C.; Zhou, Y.; Lee, J.; Gokalp, D.; Herrema, H.; Park, S.W.; Davis, R.J.; Ozcan, U. p38 MAPK-mediated regulation of Xbp1s is crucial for glucose homeostasis. Nat. Med. 2011, 17, 1251–1260. [Google Scholar] [CrossRef]
  18. Majumder, M.; Huang, C.; Snider, M.D.; Komar, A.A.; Tanaka, J.; Kaufman, R.J.; Krokowski, D.; Hatzoglou, M. A novel feedback loop regulates the response to endoplasmic reticulum stress via the cooperation of cytoplasmic splicing and mRNA translation. Mol. Cell. Biol. 2012, 32, 992–1003. [Google Scholar] [CrossRef]
  19. Navon, A.; Gatushkin, A.; Zelcbuch, L.; Shteingart, S.; Farago, M.; Hadar, R.; Tirosh, B. Direct proteasome binding and subsequent degradation of unspliced XBP-1 prevent its intracellular aggregation. FEBS Letters 2010, 584, 67–73. [Google Scholar] [CrossRef]
  20. Park, S.W.; Zhou, Y.; Lee, J.; Lu, A.; Sun, C.; Chung, J.; Ueki, K.; Ozcan, U. The regulatory subunits of PI3K, p85α and p85β, interact with XBP-1 and increase its nuclear translocation. Nat. Med. 2010, 16, 429–437. [Google Scholar] [CrossRef]
  21. Tirosh, B.; Iwakoshi, N.N.; Glimcher, L.H.; Ploegh, H.L. Rapid turnover of unspliced Xbp-1 as a factor that modulates the unfolded protein response. J. Biol. Chem. 2006, 281, 5852–5860. [Google Scholar]
  22. Wang, F.-M.; Chen, Y.-J.; Ouyang, H.-J. Regulation of unfolded protein response modulator XBP1s by acetylation and deacetylation. Biochem. J. 2011, 433, 245–252. [Google Scholar] [CrossRef]
  23. Winnay, J.N.; Boucher, J.; Mori, M.A.; Ueki, K.; Kahn, C.R. A regulatory subunit of phosphoinositide 3-kinase increases the nuclear accumulation of X-box-binding protein-1 to modulate the unfolded protein response. Nat. Med. 2010, 16, 438–445. [Google Scholar] [CrossRef]
  24. Yanagitani, K.; Imagawa, Y.; Iwawaki, T.; Hosoda, A.; Saito, M.; Kimata, Y.; Kohno, K. Cotranslational targeting of XBP1 protein to the membrane promotes cytoplasmic splicing of its own mRNA. Mol. Cell 2009, 34, 191–200. [Google Scholar] [CrossRef]
  25. Yanagitani, K.; Kimata, Y.; Kadokura, H.; Kohno, K. Translational pausing ensures membrane targeting and cytoplasmic splicing of XBP1u mRNA. Science 2011, 331, 586–589. [Google Scholar] [CrossRef]
  26. Yoshida, H.; Oku, M.; Suzuki, M.; Mori, K. pXBP1(u) encoded in XBP1 pre-mRNA negatively regulates unfolded protein response activator pXBP1(s) in mammalian ER stress response. J. Cell. Biol. 2006, 172, 565–575. [Google Scholar] [CrossRef]
  27. Liou, H.C.; Boothby, M.R.; Finn, P.W.; Davidon, R.; Nabavi, N.; Zeleznik-Le, N.J.; Ting, J.P.; Glimcher, L.H. A new member of the leucine zipper class of proteins that binds to the HLA DR α promoter. Science 1990, 247, 1581–1584. [Google Scholar]
  28. Glimcher, L.H. XBP1: The last two decades. Ann. Rheum. Dis. 2010, 69 (Suppl. 1), i67–i71. [Google Scholar] [CrossRef]
  29. Reimold, A.M.; Iwakoshi, N.N.; Manis, J.; Vallabhajosyula, P.; Szomolanyi-Tsuda, E.; Gravallese, E.M.; Friend, D.; Grusby, M.J.; Alt, F.; Glimcher, L.H. Plasma cell differentiation requires the transcription factor XBP-1. Nature 2001, 412, 300–307. [Google Scholar] [CrossRef]
  30. Lee, A.H.; Chu, G.C.; Iwakoshi, N.N.; Glimcher, L.H. XBP-1 is required for biogenesis of cellular secretory machinery of exocrine glands. EMBO J. 2005, 24, 4368–4380. [Google Scholar] [CrossRef]
  31. Lee, A.-H.; Heidtman, K.; Hotamisligil, G.S.; Glimcher, L.H. Dual and opposing roles of the unfolded protein response regulated by IRE1α and XBP1 in proinsulin processing and insulin secretion. Proc. Natl. Acad. Sci. USA 2011, 108, 8885–8890. [Google Scholar] [CrossRef]
  32. Lee, A.-H.; Scapa, E.F.; Cohen, D.E.; Glimcher, L.H. Regulation of hepatic lipogenesis by the transcription factor XBP1. Science 2008, 320, 1492–1496. [Google Scholar] [CrossRef]
  33. Reimold, A.M.; Etkin, A.; Clauss, I.; Perkins, A.; Friend, D.S.; Zhang, J.; Horton, H.F.; Scott, A.; Orkin, S.H.; Byrne, M.C.; et al. An essential role in liver development for transcription factor XBP-1. Genes Dev. 2000, 14, 152–157. [Google Scholar]
  34. Iwakoshi, N.N.; Pypaert, M.; Glimcher, L.H. The transcription factor XBP-1 is essential for the development and survival of dendritic cells. J. Exp. Med. 2007, 204, 2267–2275. [Google Scholar] [CrossRef]
  35. Martinon, F.; Chen, X.; Lee, A.-H.L.; Glimcher, L.H. TLR activation of the transcription factor XBP1 regulates innate immune responses in macrophages. Nat. Immunol. 2010, 11, 411–418. [Google Scholar] [CrossRef]
  36. Kaser, A.; Lee, A.-H.; Franke, A.; Glickman, J.N.; Zeissig, S.; Tilg, H.; Nieuwenhuis, E.E.S.; Higgins, D.E.; Schreiber, S.; Glimcher, L.H.; et al. XBP1 links ER stress to intestinal inflammation and confers genetic risk for human inflammatory bowel disease. Cell 2008, 134, 743–756. [Google Scholar] [CrossRef]
  37. Ozcan, U.; Cao, Q.; Yilmaz, E.; Lee, A.H.; Iwakoshi, N.N.; Ozdelen, E.; Tuncman, G.; Gorgun, C.; Glimcher, L.H.; Hotamisligil, G.S. Endoplasmic reticulum stress links obesity, insulin action, and type 2 diabetes. Science 2004, 306, 457–461. [Google Scholar] [CrossRef]
  38. Cox, J.S.; Shamu, C.E.; Walter, P. Transcriptional induction of genes encoding endoplasmic reticulum resident proteins requires a transmembrane protein kinase. Cell 1993, 73, 1197–1206. [Google Scholar] [CrossRef]
  39. Mori, K.; Ma, W.; Gething, M.J.; Sambrook, J. A transmembrane protein with a cdc2+/cdc28-related kinase activity is required for signaling from the ER to the nucleus. Cell 1993, 74, 743–756. [Google Scholar] [CrossRef]
  40. Sidrauski, C.; Walter, P. The transmembrane kinase Ire1p is a site-specific endonuclease that initiates mRNA splicing in the unfolded protein response. Cell 1997, 90, 1031–1039. [Google Scholar] [CrossRef]
  41. Tirasophon, W.; Welihinda, A.A.; Kaufman, R.J. A stress response pathway from the endoplasmic reticulum to the nucleus requires a novel bifunctional protein kinase/endoribonuclease (Ire1p) in mammalian cells. Genes Dev. 1998, 12, 1812–1824. [Google Scholar] [CrossRef]
  42. Wang, X.Z.; Harding, H.P.; Zhang, Y.; Jolicoeur, E.M.; Kuroda, M.; Ron, D. Cloning of mammalian Ire1 reveals diversity in the ER stress responses. EMBO J. 1998, 17, 5708–5717. [Google Scholar] [CrossRef]
  43. Lee, A.H.; Iwakoshi, N.N.; Glimcher, L.H. XBP-1 regulates a subset of endoplasmic reticulum resident chaperone genes in the unfolded protein response. Mol. Cell. Biol. 2003, 23, 7448–7459. [Google Scholar] [CrossRef]
  44. Shaffer, A.L.; Shapiro-Shelef, M.; Iwakoshi, N.N.; Lee, A.H.; Qian, S.B.; Zhao, H.; Yu, X.; Yang, L.; Tan, B.K.; Rosenwald, A.; et al. XBP1, downstream of Blimp-1, expands the secretory apparatus and other organelles, and increases protein synthesis in plasma cell differentiation. Immunity 2004, 21, 81–93. [Google Scholar] [CrossRef]
  45. Bartoszewski, R.; Brewer, J.W.; Rab, A.; Crossman, D.K.; Bartoszewska, S.; Kapoor, N.; Fuller, C.; Collawn, J.F.; Bebok, Z. The unfolded protein response (UPR)-activated transcription factor X-box-binding protein 1 (XBP1) induces microRNA-346 expression that targets the human antigen peptide transporter 1 (TAP1) mRNA and governs immune regulatory genes. J. Biol. Chem. 2011, 286, 41862–41870. [Google Scholar]
  46. Iwakoshi, N.N.; Lee, A.H.; Vallabhajosyula, P.; Otipoby, K.L.; Rajewsky, K.; Glimcher, L.H. Plasma cell differentiation and the unfolded protein response intersect at the transcription factor XBP-1. Nat. Immunol. 2003, 4, 321–329. [Google Scholar]
  47. Back, S.H.; Lee, K.; Vink, E.; Kaufman, R.J. Cytoplasmic IRE1α-mediated XBP1 mRNA splicing in the absence of nuclear processing and endoplasmic reticulum stress. J. Biol. Chem. 2006, 281, 18691–18706. [Google Scholar] [CrossRef]
  48. Uemura, A.; Oku, M.; Mori, K.; Yoshida, H. Unconventional splicing of XBP1 mRNA occurs in the cytoplasm during the mammalian unfolded protein response. J. Cell Sci. 2009, 122, 2877–2886. [Google Scholar] [CrossRef]
  49. Stephens, S.B.; Dodd, R.D.; Brewer, J.W.; Lager, P.J.; Keene, J.D.; Nicchitta, C.V. Stable ribosome binding to the endoplasmic reticulum enables compartment-specific regulation of mRNA translation. Mol. Biol. Cell 2005, 16, 5819–5831. [Google Scholar] [CrossRef]
  50. Duan, Q.; Wang, X.; Gong, W.; Ni, L.; Chen, C.; He, X.; Chen, F.; Yang, L.; Wang, P.; Wang, D.W. ER stress negatively modulates the expression of the mir-199a/214 cluster to regulate tumor survival and progression in human hepatocellular cancer. PLoS One 2012, 7, e31518. [Google Scholar]
  51. Bartel, D.P. MicroRNAs: target recognition and regulatory functions. Cell 2009, 136, 215–233. [Google Scholar] [CrossRef]
  52. Behrman, S.; Acosta-Alvear, D.; Walter, P. A CHOP-regulated microRNA controls rhodopsin expression. J. Cell Biol. 2011, 192, 919–927. [Google Scholar] [CrossRef]
  53. Deng, J.; Lu, P.D.; Zhang, Y.; Scheuner, D.; Kaufman, R.J.; Sonenberg, N.; Harding, H.P.; Ron, D. Translational repression mediates activation of nuclear factor kappa B by phosphorylated translation initiation factor 2. Mol. Cell. Biol. 2004, 24, 10161–10168. [Google Scholar] [CrossRef]
  54. Jiang, H.Y.; Wek, S.A.; McGrath, B.C.; Scheuner, D.; Kaufman, R.J.; Cavener, D.R.; Wek, R.C. Phosphorylation of the alpha subunit of eukaryotic initiation factor 2 is required for activation of NF-kappaB in response to diverse cellular stresses. Mol. Cell. Biol. 2003, 23, 5651–5663. [Google Scholar] [CrossRef]
  55. Romero-Ramirez, L.; Cao, H.; Nelson, D.; Hammond, E.; Lee, A.-H.; Yoshida, H.; Mori, K.; Glimcher, L.H.; Denko, N.C.; Giaccia, A.J.; et al. XBP1 is essential for survival under hypoxic conditions and is required for tumor growth. Cancer Res. 2004, 64, 5943–5947. [Google Scholar] [CrossRef]
  56. Hay, R.T. SUMO: A history of modification. Mol. Cell 2005, 18, 1–12. [Google Scholar] [CrossRef]
  57. Lee, A.-H.; Iwakoshi, N.N.; Anderson, K.C.; Glimcher, L.H. Proteasome inhibitors disrupt the unfolded response in myeloma cells. Proc. Natl. Acad. Sci. 2003, 100, 9946–9951. [Google Scholar]
  58. Zhou, Y.; Lee, J.; Reno, C.M.; Sun, C.; Park, S.W.; Chung, J.; Lee, J.; Fisher, S.J.; White, M.F.; Biddinger, S.B.; et al. Regulation of glucose homeostasis through a XBP-1-FoxO1 interaction. Nat. Med. 2011, 17, 356–365. [Google Scholar] [CrossRef]
  59. Accili, D.; Arden, K.C. FoxOs at the crossroads of cellular metabolism, differentiation, and transformation. Cell 2004, 117, 421–426. [Google Scholar] [CrossRef]
  60. Gross, D.N.; van den Heuvel, A.P.J.; Birnbaum, M.J. The role of FoxO in the regulation of metabolism. Oncogene 2008, 27, 2320–2336. [Google Scholar] [CrossRef]
  61. Yoshida, H.; Uemura, A.; Mori, K. pXBP1(u), a negative regulator of the unfolded protein response activator pXBP1(s), targets ATF6 but not ATF4 in proteasome-mediated degradation. Cell Struct. Funct. 2009, 34, 1–10. [Google Scholar] [CrossRef]
  62. Newman, J.R.S.; Keating, A.E. Comprehensive identification of human bZIP interactions with coiled-coil arrays. Science 2003, 300, 2097–2101. [Google Scholar] [CrossRef]
  63. Hotamisligil, G.S. Endoplasmic reticulum stress and the inflammatory basis of metabolic disease. Cell 2010, 140, 900–917. [Google Scholar] [CrossRef]
  64. Koong, A.C.; Chauhan, V.; Romero-Ramirez, L. Targeting XBP-1 as a novel anti-cancer strategy. Cancer Biol. Ther. 2006, 5, 756–759. [Google Scholar] [CrossRef]
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