The Road Well Traveled: From Inflammasomes to Collagen Export During Fibrosis

Abstract

Collagen export from the endoplasmic reticulum is required for normal tissue homeostasis, and yet, in fibrotic disorders, this process is significantly upregulated. In this review, we will focus on the signaling cascade from the inflammasome and how that promotes collagen via proinflammatory/profibrotic cytokines. Concordantly, these cytokines also induce the expression of TANGO1 to cope with the increased movement of collagen through the endoplasmic reticulum. In normal and fibrotic cells, this pathway is finely tuned to meet the necessary demand in collagen export. Currently, the role of TANGO1 in fibrotic disorders and how the inflammasome induces its expression is not well understood. In this review, we will assimilate the current information concerning inflammasome activation and how it induces TANGO1 expression, leading to fibrosis.

1. Introduction

Collagen is the most abundant protein in mammals, and collagen synthesis has been extensively studied in normal and fibrotic conditions. Collagen must be correctly folded in the endoplasmic reticulum (ER) before it is exported from the ER to the Golgi. An essential protein for collagen folding and export is HSP47 [1]. HSP47 binds to the collagen triple helix and facilitates the correct folding of the protein. HSP47 also interacts with TANGO1 (Transport and Golgi Organization 1) [2], recruiting the collagen molecules into the growing COPII vesicle for export [3]. HSP47 is found in cells that express collagen [4], and its expression directly correlates with collagen expression [5]. Once in the Golgi, collagen undergoes further modifications, such as glycosylation, which helps stabilize the triple helix [6], and then it is secreted and further modified to form the mature fibers [7].

Fibrosis, also known as fibrotic scarring, is a pathological process that, if left unaddressed, can lead to tissue stiffening and the failure of the organ to function due to the excessive deposition of collagen and other ECM proteins in the tissues [8,9]. Fibrosis can be localized to a single organ/tissue [10,11,12] or be systemic, affecting multiple organs and tissues [13,14,15]. Depending on the affected organ, fibrosis can cause significant morbidity and can be fatal. Although the mechanism of fibrosis is similar to wound healing in many ways [16], fibrosis involves the uncontrolled deposition of collagen and other extracellular matrix proteins in the tissues. This contrasts with wound healing, where collagen secretion is controlled and stops once the wound has closed [17,18,19]. Currently, it is not fully understood why fibrosis occurs. Still, diverse factors, including viruses [20,21], bacteria [22], environmental insults [23,24,25,26], genetic predisposition [27,28], autoimmune diseases [29,30,31], and cancer [32,33,34,35], can all contribute to the development of fibrosis. Age is one of the most significant risk factors for developing fibrosis [36,37]. The export of large proteins such as collagen from the ER under basal conditions has been well studied in Drosophila [38] and various cell lines [39,40], and the role of TANGO1 in this process is well known. However, little is known about TANGO1 in the context of fibrosis. The current information in collagen export and the role of the inflammasome intersecting with TANGO1 expression in this process is discussed below.

2. Systemic Sclerosis

Systemic sclerosis (scleroderma, SSc) is a fibrotic disease with autoimmune and vascular components [41,42,43]. It manifests with progressive skin [44] and internal organ fibrosis [45]. Its clinical forms are complex and heterogeneous, with presentations ranging from limited/restricted skin involvement (LcSSc) to widespread diffuse skin involvement (DcSSc) [46]. DcSSc results in an extensive cutaneous presentation with severe internal organ involvement that starts early in the disease [46]. Like the other fibrotic disorders, age [47,48], race [47], genetics [49,50], epigenetic changes [51], genetic mutations [52] and environmental factors [53,54,55] influence the development and progression of SSc. The most potent genetic risk factor is being female, with about eight females to every one male diagnosed with SSc [56,57]. The etiology of SSc is still unknown, and no single unifying hypothesis explains all aspects of disease pathogenesis.

3. Fibroblasts

Fibroblasts are sentinel cells [58] but play a significant role in maintaining the integrity of tissues. They are crucial for forming and supporting the structure of the connective framework within the skin and internal organs. They are the primary cells that produce the extracellular matrix, composed primarily of collagens, elastin, and glycosaminoglycans. These proteins provide strength, elasticity, and support to the organs and tissues. An added function of fibroblasts is to change the extracellular matrix as they mediate the cross-linking of the collagens and, therefore, maturation. Fibroblasts sense the microenvironment, react to it, and can become activated [59,60]. Activated fibroblasts differentiate into myofibroblasts that produce excessive amounts of collagen to aid wound repair or, in pathologic conditions, cause fibrosis. Like any sentinel cell in the body, they have functional inflammasomes that, when activated, promote collagen deposition in wound healing or fibrosis.

A hallmark of myofibroblasts is the cell’s increased expression of stress fibers [61,62] caused by their response to various inflammatory cytokines. Alpha-smooth muscle actin is elevated in myofibroblasts [63]. One of the functions of α-smooth muscle actin is to connect focal adhesions at the cell surface to tropomyosins spread throughout the cell [64], allowing the cell to migrate through the tissues by pulling itself forward [65]. Fibroblast migration is essential in wound healing [66]; however, it also plays a significant role in fibrosis [67,68,69]. The recruitment of fibroblasts to the wound site is thought to occur by cytokines released during wounding or inflammation [70]. These processes activate and are driven by the inflammasomes that release and attract cytokines fibroblasts [71,72]. Fibroblasts/myofibroblasts must have a motile function to traverse various tissue microenvironments, and they are required to degrade, repair, or remodel the extracellular matrix [73,74].

4. NLRP3 Inflammasome Activity in Fibrotic Diseases

Inflammasomes rarely act alone, and many are often upregulated during fibrosis [75]. This makes studying the contribution of various inflammasomes complex. However, careful analyses using in vitro studies and animal models have identified the ability of inflammasomes to drive fibrosis.

The NLRP3 inflammasome platform is strongly associated with fibrosis and is activated in SSc. The nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasome is the most extensively studied of the inflammasome platforms, and its activity is highly associated with diverse forms of fibrosis. NLRP3 has been associated with fibrosis in many organs, including the liver [48,49], heart [50], kidney [51], lung [46,52] and skin [46]. Bleomycin sulfate is an antibiotic chemotherapeutic drug often used to induce collagen expression in vitro and in vivo. It damages DNA and activates the NLRP3 inflammasome. Using bleomycin, numerous studies have confirmed the role of NLRP3 in skin and lung fibrosis. This molecule has significantly contributed to understanding this inflammatory platform’s pathogenic mechanisms. Indeed, inhibiting caspase-1 with YVAD ameliorates fibrosis in animals and in vitro studies, confirming the inflammasome’s role in fibrosis.

NLRP3 can be activated in a wide variety of different cells by a wide variety of diverse triggers started by microbe-derived pathogen-associated molecular patterns (called PAMPs) [76], danger-associated molecular patterns (called DAMPs) [77], or homeostasis-altering molecular processes (called HAMPs) [78] that are generated by the host cell. It is believed that many of the different stimuli causing NLRP3 activation converge on changes in ion flux [79], changes in metabolic flux [80], or dysfunction in cellular organelles [81] (Figure 1). It is made up of seven NLRP3 molecules that interact in a wheel-like structure and position inactive procaspase-1 in a manner where they can autocatalytically cleave themselves. Once activated, caspase-1 triggers the cleavage of the proinflammatory cytokines IL-1β and IL-18. Both these cytokines, synthesized as biologically inactive precursors, require processing for their activation by caspase-1, then IL-1β and IL-18 start and perpetuate an inflammatory response. The function of IL-1β is to amplify inflammation by recruiting immune cells to the site of the infection or damage. It also regulates the adaptive immune response [82] and stimulates the hypothalamus to induce fever [83]. IL-18 is essential in the production of IFN-γ [84]. It induces the cytolytic activity of NK cells and activates T cells [85]. The secretome mediated by caspase-1 releases numerous other proteins involved in inflammation, cell protection, and tissue repair cascades [86]. Inflammasomes were first studied in immune cells [87,88]. However, it is now well recognized that they are found in many other types of cells, including stromal and mesenchymal cells, such as epithelial cells [71,89], keratinocytes [90,91] and fibroblasts [72] and hepatic stellate cells [92].

In the right setting, IL-1β is profibrotic. IL-1β increases leukocyte recruitment and induces other profibrotic mediators such as IL-6 [93,94,95]. It can upregulate TGF-β1 [96] and is increased in SSc [97,98,99]. IL-1 promotes the fibrogenic macrophage phenotype that secretes TGF-β1 [100]. IL-1 added to fibroblasts can directly enhance TGF-β1 expression and, therefore, can drive matrix production [96], and moreover, it has been shown to promote fibroblast proliferation [101], also contributing to the fibrotic process. Additional downstream signaling by IL-1β can promote α-smooth muscle actin (α-SMA) expression [72]. Studies have found a more significant response to IL-1 in fibroblasts derived from fibrotic tissues [102] leading to enhanced myofibroblast development in these tissues. This suggests that there could be increased numbers of the IL-1 receptor in these cells, but this has not been tested. Although less studied in the fibrotic context, it has been shown that IL-18 is also profibrotic [103,104].

In the normal context of wound healing, IL-1 naturally downregulates TGF-β1 once the wound is closed. However, the mechanism behind fibrosis is complex and poorly understood, and there is no visible “wound”. Serum IL-1β levels are not reliable indicators for the role of this cytokine in fibrosis or in many inflammatory diseases [105] as the effects can be localized. However, relevant to fibrosis is that long-term exposure to IL-1β enhances TGF-β1 signaling [106]. Overall, the data suggest that IL-1β has dual signaling capabilities.

Once NLRP3 is activated, TGF-β1 is induced as a downstream mediator of this process [107] due to IL-1 receptor and IL-18 receptor signaling, leading to increased collagen expression and deposition into the tissues. This increased collagen expression places the ER under immense strain to export collagen quickly and efficiently [108,109]. Furthermore, the activation of NLRP3 contributes to the development of myofibroblasts [72]. In our studies, we saw that the inhibition of caspase-1 in SSc myofibroblasts decreased α-smooth muscle actin stress fibers and total protein. In addition, it appeared that the cells were dedifferentiating back to quiescent fibroblasts [72]. Nguyen and colleagues showed that the activation of NLRP3 was driving the myofibroblast phenotype and collagen deposition [110]. When they inactivated NLRP3 with MCC950 or inhibited caspase-1 with YVAD, collagen secretion was reduced.

One intriguing observation about fibrosis is that older individuals are more susceptible to it. Stout-Delgaldo and colleagues were able to elucidate why. They showed an increased activation of NLRP3 in aged mice, while younger mice had less activation. Furthermore, they found pro-IL-1β and pro-IL-18 expression to be substantially higher in the aged mice, and they speculated that this was because of enhanced NF-kB caused by increased cell damage and oxidative stress [111]. They also found an increased expression of P2X7 receptors in the aged cells, and they speculated that this could further amplify NLRP3 inflammasome signaling and elevate inflammatory mediators [111], suggesting an autocrine loop. Studies by Riteau and colleagues further defined this. They showed that extracellular ATP was a danger signal and activated the P2X7 receptor, driving lung inflammation and fibrosis [112]. SSc tends to be a disease with the onset clustering in individuals between 30 and 50 years of age [113], although children do obtain SSc [114]. The onset of SSc can also occur much later than the general norm [48].

5. Post-Translational Modifications of NLRP3

NLRP3 has three post-translational modifications that regulate its activity: ubiquitylation, sumoylation, and phosphorylation. They have been well studied but not in the context of fibrosis. In its inactive state, NLRP3 is ubiquitylated and must be deubiquitylated to become active [115]. Several proteins regulate the ubiquitylation and deubiquitylation of NLRP3. The F-box and the leucine-rich repeat protein-2 recognize the tryptophan residue at position 73 on NLRP3, which targets the lysine-689 residue for ubiquitylation and subsequent protein degradation [116]. LPS increases the half-life of NLRP3 because F-box and leucine-rich repeat protein-2 are degraded [116]. Other factors also control the activated/inactivated state of NLRP3. NLRP3 exists in cells in an inactivated state, which is regulated in part by cAMP. NLRP3 becomes active when cAMP disengages from the protein. [117]. TRIM31 encodes the protein E3 ubiquitin–protein ligase, which is crucial for regulating NLRP3 activity. TRIM31 causes the ubiquitylation and the proteasomal degradation of NLRP3 [118], and as such, it is a negative regulator of inflammasome activity [119]. There are limited studies, but it has been shown that NLRP3 ubiquitylation is associated with fibrosis. Liver inflammation due to hepatitis C infections can ultimately lead to liver cirrhosis and fibrosis. Due to the infection, NLRP3 is activated in hepatitis C virus-infected hepatocytes, and the resulting deubiquitylation of NLRP3 causes a chronic increase in inflammation. These observations were confirmed when deubiquitinases were used to prevent NLRP3 activation. As a downstream result of this inhibition, there was reduced IL-1β secretion. An unexpected observation of this study was a decrease in viral protein and a corresponding reduction in the release of the hepatitis C virus from the cells [120].

The SUMOylation of NLRP3 also plays a role in the activity of this protein. SUMOylation is a post-transcriptional modification that tags various residues with a small ubiquitin-like modifier (SUMO). The SUMO E3-ligase MAPL SUMOylates NLPR3, and this renders the protein inactive [121]. For NLRP3 to be activated, the SUMOylation tags have to be removed by sentrin-specific protease-6 and -7. If there is a deficiency in these proteases due to the downregulation or deletion of the specific genes, NLRP3 activation is reduced [121]. In contrast, to further complicate the role of SUMOylation in the regulation of NLRP3 activity, the SUMO1 protein catalyzes the SUMOylation of NLRP3, aiding in its activation, but sentrin-specific protease-3 removes this SUMOylation, causing deactivation [122]. Other changes are necessary for NLRP3 activity, and they include the phosphorylation of various residues [123], while the phosphorylation of alternate residues is associated with NLRP3 inactivation [124,125].

6. TANGO1 Drives ER to Golgi Trafficking of Collagen Export During Fibrosis

The export of collagen out of the ER was once considered a size paradox. Still, it is now recognized that TANGO1, which is coded by the MIA3 (MIA SH3 Domain ER Export Factor 3) gene, creates tunnels along which collagen travels to the Golgi. TANGO1 resides in ER exit sites and interacts with collagen via its HSP47 binding site. Both TANGO1 and HSP47 are necessary proteins for collagen export. Saito and colleagues [38] reported the functional role of TANGO1 in orchestrating collagen loading into the tubes. Other studies by Saito and colleagues [126] further demonstrated that TANGO1 organized the ER exit sites, a necessary step in collagen export. Overall, the scientific community is only now starting to appreciate the complexity of collagen export.

Maiers and colleagues [127] were the first to report that elevated TANGO1 was required for fibrosis. They showed in hepatic stellate cells increased TANGO1 protein. They then proved that the deletion of TANGO1 blocked the secretion of type I collagen. The deletion of TANGO1 led to the retention of procollagen in the ER, resulting in stress and the unfolded protein response. They further showed that the unfolded protein response induced TANGO1 by increasing TGF-β1 via the X-box binding protein 1 transcription factor [127].

In our studies, we show that TANGO1 is elevated SSc fibroblasts [128]. SSc lesions have increased numbers of secretory myofibroblasts, and to meet the increased secretory demand, myofibroblasts must depend on the ER to process and export the augmented protein load and, therefore, must have elevated TANGO1 protein levels. Using an antibody that targets the two predominant splice variants of TANGO1, we show that TANGO1-Long and TANGO1-Short were concurrently elevated in SSc fibroblasts [128].

As our prior studies showed the dependency of fibrosis on inflammasome activity and caspase-1, we wondered if TANGO1 expression was dependent on this inflammatory cascade. Using the caspase-1 chemical inhibitor, Z-YVAD-FMK, we were able to show a decreased expression of TANGO1-Long and TANGO1-Short. This observation strongly suggests the involvement of the inflammasome in the expression of TANGO1. Taking this observation further and looking at downstream cytokine signaling, we blocked the TGF-β receptor with the ALK5 inhibitor and showed decreased total collagen, type I collagen, TANGO1-Long, and TANGO1-Short. We believe that this effect on TANGO1 was primarily via decreased collagen. Because less collagen was trafficked through the ER, this took pressure off the collagen export system. This meant there was less need for TANGO1, and its expression decreased. We then determined if IL-1 was involved in TANGO1 expression, and we used an IL-1 receptor antagonist to block the receptor. We found that TANGO1-Long and TANGO1-Short were lowered, but collagen was not decreased and kept in the cell. In those studies, we did not figure out if there was ER stress and an unfolded protein response with the retained collagen. The net effect of inhibiting both the TGF-β receptor and the IL-1 receptor was significantly reduced collagen secretion and, therefore, less fibrosis [128]. Further defining the role of TANGO1 in total protein secretion in SSc fibroblasts, we examined the culture media from the fibroblasts treated with the caspase-1 inhibitor, the ALK5 inhibitor, and the IL-1 receptor antagonist. We found that high-molecular-weight proteins were significantly affected by the inhibitors, and there was less total protein in the culture media that had a molecular weight above the arbitrary cut-off of 100 kDa [128].

Elegant studies by Raote and colleagues [129] and more recently by Reynolds and colleagues [130] show that TANGO1 assembles into rings, controlling the formation of the COPII docking sites for protein export. Further, the over-expression of the cytoplasmic domain of TANGO1 can increase the size and density of ER exit sites and can also increase the number of Golgi [130]. Overall, this suggests that TANGO1 can regulate both ER exit sites and the Golgi, although more studies are needed to confirm this observation. It is also likely that the flexibility of the COPII coat during protein export is dependent on the protein burden [131]. Some data suggest that procollagen export can occur directly from the ER to the Golgi in the absence of COPII carriers [132]; however, this seems unlikely as cTAGE5/TANGO1 interacts with the COPII protein Sec23.

More recent studies have shown that ER exit sites are segregated (Figure 2). The sorting and concentration of cargo molecules at ER exit sites occur before they exit the ER [133]. Saxena and colleagues [134] found that the binding between TANGO1-S and Sec23A stalls the exit of protein (ERGIC53-GFP was used as a marker), and ERGIC53-GFP accumulates at the ER exit site. It has been hypothesized that the stable capture of Sec23/Sec24 by TANGO1 immobilizes the associated cargo receptors, with their cargo inhibiting their exit and, at the same time, allowing for the necessary membrane remodeling and fission for larger cargoes [134]. Intriguingly, the collagen protein is concentrated at approximately half of the ER exit sites, while smaller cargoes accumulate in all exit sites. The authors also suggest that the interaction between Sec23/Sec24 and TANGO1-S acts like a “kinetic brake” that the cell uses to generate appropriate export vesicles required for the size of the cargo [134]. When there is a depletion of TANGO1, or when Sec23A traps TANGO1, all ER exit sites are affected, and there is a global defect in protein secretion. However, Saxena et al. hypothesized that the segregation of ER exit sites provides a way for the ER to build tunneling for collagen export while keeping other ER exit sites open for the trafficking of smaller cargo [134]. Our studies also support this observation. We found that when TANGO1 expression was reduced with YVAD, IL-1RA, or ALK5, the larger cargoes were significantly affected, while the export of smaller cargos remained largely unchanged [128].

7. Activation of TANGO1 by O-Glycosylation

The ability of TANGO1 to function in the export of collagen out of the ER is not just dependent on its expression. Secretion is critical in many tissues, for example, the gut, and O-glycosylation plays a role in this process [135]. The loss of O-glycosylation disrupts secretion, further confirming its importance [136,137]. Intriguingly, the loss of O-glycosylation resulted in decreased tiggrin release [138]. Tiggrin is a large extracellular matrix protein requiring TANGO1 for its export [139]. Considering these observations, Zhang and colleagues studied the influence of O-glycosylation on secretion and secretory vesical size in the ER and found that O-glycosylation was critical in TANGO1 activity [140]. These studies in fruit flies proved that the PGANT4 protein (Polypeptide N-acetylgalactosaminyltransferase-4), an O-glycosyltransferase, regulates TANGO1 stability and function. The O-glycans present in the TANGO1 protein were found to protect it from furin-mediated protein cleavage [140].

8. TANGO1 Mutations Correlate with Collagenopathies

While increased TANGO1 has been associated with SSc and hepatic fibrosis, mutations in the TANGO1 gene have been associated with other collagenopathies. However, classic mutations have been found in the MIA3 gene, affecting the function of TANGO1 and altering the export of collagen. These mutations are related to various collagenopathies in humans and other animals and implicate the significant role that TANGO1 has in collagen export. Studies examining the loss of TANGO1 in zebrafish found that the fish were significantly shorter in length than their wild-type counterparts and had craniofacial defects. The fish failed to thrive, and they did not survive to adulthood [141]. TANGO1 directly interacts with cTAGE5 (cutaneous T-cell lymphoma-associated antigen 5), and this interaction is necessary for large protein export. Zebrafish depleted of cTAGE5, like the TANGO1-KOs, also have a shortened stature with less collagen in the tissues, but they have better survival rates to 12 months of age [141]. This study confirms that the function of TANGO1 with one of its binding partners is needed for effective collagen secretion.

Similar pathologies are seen in mice lacking MIA3. Without TANGO1, the fetal pups showed chondrodysplasia, dwarfing, and tissue fragility. They exhibited a shortening of the snout and limbs, and death was due to the absence of an ossified skeleton [142]. Cane Corso dogs have dental, skeletal, and retinal abnormalities, and only recently, they were found to have mutations in the MIA3 gene. Dogs have brittle, discolored, and translucent teeth. They have disproportionate growth, being smaller than their normal littermates. They have progressive retinal degeneration, resulting in vision loss as they age. Cane Corso dogs have been identified with a splice mutation leading to the skipping of two exons in the MIA3 gene. This results in a truncated TANGO1 protein, but it does have some residual activity, allowing for some collagen to be exported and for the animals to survive to adulthood [143]. The mutation in the MIA3 gene is an autosomal recessive disease, and carriers show no phenotypic effects. This suggests that one functional copy of TANGO1 is sufficient for normal procollagen export.

To date, only a few cases have identified TANGO1 mutations in humans. Ehler’s Danlos syndrome typically occurs with classic mutations in connective tissue genes. However, one study investigated patients diagnosed with Ehlers–Danlos syndrome who were negative for these mutations and identified four mutations and one frameshift mutation in the MIA3 gene [144]. All mutations were heterozygous and pathogenic. The patient carrying truncated TANGO1 displayed skeletal abnormalities such as scoliosis, osteopenia, brachydactyly and clinodactyly, and dentinogenesis imperfecta. They also had a mild intellectual disability. A further study of this individual showed a substantial reduction in COL1A1 secretion [144].

Two consanguineous families point to the importance of TANGO1. In one consanguineous family, the fetuses presented with early lethality due to the complete absence of bone formation. This was due to a homozygous frameshift in the MIA3 gene that caused a premature termination codon and a lack of the TANGO1 protein [145]. In the other family, there was a homozygous mutation that led to exon eight skipping in the MIA3 gene, resulting in the formation of a truncated TANGO1 protein affecting collagen secretion [146]. Four homozygous mild-to-moderately affected sons showed similarities with the Cane Corso dogs, demonstrating dentinogenesis imperfecta resulting in the delayed eruption of permanent teeth and numerous other skeletal abnormalities. They also had a mild intellectual disability [146]. An intriguing observation about this family is the mild intellectual disability in the affected individuals, which begs the following question: is TANGO1 required for the secretion of large proteins that aid the normal functioning of the brain? Collagen and extracellular matrix proteins are found in the brain, but three other large proteins (>130 kDa) are also secreted. These include versican, brevican, and neurocan, all detected in the cortex. Intriguingly, mutations in the TANGO1 binding partner cTAGE5 also result in abnormal changes in the brain. These changes result in hypomyelination, compromised synaptic functionality, and aberrant behaviors in mice [147,148].

9. Targeting TANGO1 to Treat Fibrosis

At best, treating fibrotic disorders has been very difficult, and many small molecules fail clinical trials. This could be because there are many ways to induce collagen expression. Targeting various cytokines has shown some promise in slowing disease progression; however, none have successfully stopped disease progression [149,150]. Being able to control the amount of collagen secreted during fibrosis would benefit a spectrum of diseases that are very difficult to control. Therefore, targeting TANGO1 and preventing collagen export is an attractive idea. Intriguingly, the depletion of TANGO1 causes the loss of its binding partner, cTAGE5 [151]. At ER exit sites, TANGO1 and cTAGE5 have complementary functions; TANGO1 recruits the collagen protein, while cTAGE5 helps to build the export vehicle [152,153]. The interaction between these two proteins is highly stable. A study showing peptides that bound to the CC2 regions on TANGO1 and cTAGE5 inhibited the interaction between these two proteins and reduced collagen secretion [154]. Intriguingly, Raote and colleagues found that the inhibition between TANGO1 and cTAGE5 resulted in a minor collagen accumulation in the ER. They suggest that the accumulated collagen could have been cleared by autophagy, ER-phage, and/or proteasomal degradation [154]. However, studies have yet to confirm this observation. In any event, these findings show immense promise for the development of therapeutics that target the function of TANGO1 in the treatment of fibrotic disorders.

10. Discussion

This review aimed to assimilate the current information that outlines the role of the inflammasome in fibrosis and how this inflammatory platform induces TANGO1 and collagen expression to drive increased procollagen export from the ER. TANGO1 is an essential protein in large molecule export out of the ER and has been shown to contribute to fibrotic disorders. We are only now starting to understand the overall function of TANGO1 in exporting large proteins. Under basal conditions, collagen is secreted from fibroblasts, and there must be a baseline for the expression of TANGO1-S and TANGO1-L [128]. However, when the inflammasome is activated, there is an upregulation of procollagen, and to meet the demand for increased procollagen export from the ER, there has to be a corresponding increase in the proteins involved in the machinery that exports procollagen from the ER (Figure 3) [128]. Although many different inflammasome inhibitors have been identified, none have been approved by the FDA for the treatment of fibrosis. This suggests that targeting the inflammasome may be more complex than thought. Therefore, a greater understanding of the role of TANGO1 in the export of collagen during fibrosis is needed as this could find critical molecules that could interrupt this process and abrogate fibrosis. Furthermore, more studies will allow us to fully comprehend the pathogenic contribution that TANGO1 plays in this difficult-to-treat pathology in SSc.

11. Conclusions

TANGO1 is a critical protein involved in collagen export and was shown to be induced in fibrotic diseases. Little is known about what regulates its function during fibrosis; however, the proteins that interact with it to control its function are just starting to be elucidated. While only two TANGO1 isoforms have been investigated in fibrosis, the gene has fourteen transcript variants. This suggests that more complex interactions between the TANGO1 isoforms and the export of collagen and possibly other large cargoes could exist. Whether these other variants are relevant to fibrosis is unknown and warrants investigation. Since preventing TANGO1 from binding to cTAGE5 reduced collagen export and our studies showed reduced TANGO1 and collagen export when the inflammasome was inhibited, targeting TANGO1 could be a viable approach to treating fibrotic disorders.