The role of cellular prion protein in immune system

Abstract

Numerous studies have investigated the cellular prion protein (PrP^C) since its discovery. These investigations have explained that its structure is predominantly composed of alpha helices and short beta sheet segments, and when its abnormal scrapie isoform (PrP^Sc) is infected, PrP^Sc transforms the PrP^C, leading to prion diseases, including Creutzfeldt-Jakob disease in humans and bovine spongiform encephalopathy in cattle. Given its ubiquitous distribution across a variety of cellular types, the PrP^C manifests a diverse range of biological functions, including cell-cell adhesion, neuroprotection, signalings, and oxidative stress response. PrP^C is also expressed in immune tissues, and its functions in these tissues include the activation of immune cells and the formation of secondary lymphoid tissues, such as the spleen and lymph nodes. Moreover, high expression of PrP^C in immune cells plays a crucial role in the pathogenesis of prion diseases. In addition, it affects inflammation and the development and progression of cancer via various mechanisms. In this review, we discuss the studies on the role of PrP^C from various immunological perspectives.

INTRODUCTION

The cellular prion protein (PrP^C) is predominantly found in the brain tissue and neurons, where it is attached to the cell membrane (1-3). PrP^C is a single polypeptide composed of 208 amino acids in mice and 209 amino acids in humans (4-6). In the human PrP^C, there are two N-glycosylation sites located at residue 181 and 197. Through Western blotting, diglycosylated (36 kDa), monoglycosylated (33 kDa), and unglycosylated cerebral PrP isoforms (27 kDa) can be distinguished (7, 8). PrP^C exhibits a predominantly alpha-helical structure with a flexible N-terminal domain and a globular C-terminal domain. The C-terminal region contains three alpha helices and two short segments of the beta sheet structure. For its abnormal scrapie isoform (PrP^Sc), PrP^C undergoes a conformational change that converts alpha helices into beta-sheets (6, 9, 10). This conformational alteration is associated with the pathogenicity and transmission of prion diseases (3, 11-13). Prion diseases, also known as transmissible spongiform encephalopathies, are rare and fatal neurodegenerative disorders that are caused by the accumulation of abnormally folded PrP^Sc in the brain of humans and animals (14-16). The abnormal PrP^Sc form acts as a template and induces the conversion of PrP^C into a disease-associated form. This results in progressive accumulation of PrP^Sc, which disrupts normal brain function and causes neuronal damage (17-19). Based on the specific type and region of the brain affected, prion diseases manifest with different symptoms, including cognitive impairment, memory loss, behavioral changes, movement disorders, and severe neurological dysfunction (20, 21). Several prion diseases affect humans, including Creutzfeldt-Jakob disease (CJD), variant CJD, Gerstmann-Sträussler-Scheinker syndrome, and fatal familial insomnia (22, 23). Each disease has distinct clinical and pathological features.

PrP^C plays multifaceted roles in various cellular processes, including cell-cell adhesion (24), neuroprotection (25, 26), intracellular signaling (27), cell death and survival (28), and oxidative stress response (29). In addition to nerve cells, PrP^C is also expressed in various other cells, especially leukocytes, which are a part of the immune system (30). Despite the high expression of PrP^C in leukocytes, there have been limited scientific investigations of its precise role of PrP^C.

Here, we provide an overview of the studies investigating the role of PrP^C in the immune system. Specifically, we discuss the expression of PrP^C in immune cells and its function in the structural formation of secondary lymphoid tissues, such as the spleen and lymph nodes. Furthermore, we focused on the mechanisms of peripheral PrP^Sc replication and neuroinvasion of PrP^Sc via the immune system. We also examined the involvement of PrP^C in inflammation, cancer development, and metastasis.

EXPRESSION OF PrP^C ON IMMUNE CELLS

PrP^C is widely expressed on cell surfaces at different levels in immune cells, such as T lymphocytes (31, 32), natural killer (NK) cells (33, 34), macrophages (35), dendritic cells (DCs) (36), regulatory T cells (37) and follicular dendritic cells (FDCs) (38). Although the role of PrP^C in immune cells is not well understood, PrP^C expression increases during NK cell differentiation and functional maturation (33, 34). In addition, PrP^C in human T cells interacts with the transducer protein zeta-chain associated protein-70, which plays a critical role in the signaling pathway leading to T cell activation (31). Elevated levels of PrP^C expression have been found to facilitate T lymphocyte activation, promote cell proliferation, and enhance differentiation through the T cell receptor signaling pathway (32). DCs also exhibit upregulation of PrP^C expression after maturation (36). PrP^C also affects the phagocytic capacity of macrophages by activating the ERK1/2 and Akt kinases (35). Taken together, these studies demonstrate a correlation between the activation of immune cells and PrP^C expression, although the exact role of upregulated PrP^C remains uncertain.

Regulatory T cells, known for their ability to suppress immune responses, express higher levels of PrP^C than conventional T cells (37). Our recent study showed that PrP^C is involved in the development and function of regulatory T cells and it will be discussed in detail in section ‘Role of regulatory T cells in cancer and their relationship with PrP^C.’

FDCs are a specialized type of non-hematopoietic immune cell found primarily within the B cell area of secondary lymphoid tissues and are recognized for their high expression of PrP^C. During infection with PrP^Sc, they serve as the initial sites of accumulation in the lymphoid tissues before PrP^Sc subsequently spreads to the central nervous system (CNS) (38). However, specific ablation of PrP^C expression in FDCs using Cre-mediated recombination did not affect the normal function of FDCs (39). Further studies are required to elucidate the functions of PrP^C in FDCs.

INVOLVEMENT OF PrP^C IN THE STRUCTURE OF SECONDARY LYMPHOID TISSUES

To investigate the role of PrP^C in the structure and organization of secondary lymphoid tissues, such as the spleen and lymph nodes, several studies have been performed using mice lacking PrP^C (Prnp^0/0) and mice infected with the mouse-adapted scrapie strain ME7 (40-42). These studies revealed that PrP^C plays an important role in the formation and maintenance of secondary lymphoid tissue structures. Spleen obtained from Prnp^0/0 and ME7-infected mice showed impaired structures, with a lack of segregation between the T and B zones in the white pulp region. In both cases, there was no or significant reduction in the size of the T-zone. This can be attributed to the decreased expression of T-cell homing chemokines CCL19 and CCL21, which are involved in T-zone formation (43), leading to impaired recruitment of CD4 T cells in both mouse models (40, 41). Although both Prnp^0/0 and ME7-infected spleens exhibited impaired T-zone structures, when compared to uninfected wild-type mice, the number of lymphoid tissue inducer (LTi) cells, which are important for secondary lymphoid tissue development (44, 45) decreased in Prnp^0/0 spleens but remained unchanged in ME7-infected spleens. These results suggest that persistent PrP^C, without conversion to PrP^Sc in ME7-infected mice likely regulates both the quantity of LTi cells and their migration to the spleen.

PATHOGENESIS OF PRION DISEASES VIA IMMUNE SYSTEM

Prion diseases develop when PrP^Sc infects the host. Infected PrP^Sc continuously converts PrP^C to PrP^Sc, leading to the accumulation of PrP^Sc. Ultimately, the accumulation of PrP^Sc leads to the onset of prion diseases (46, 47). PrP^Sc accumulation primarily occurs in the immune system as immune cells express PrP^C (47-49). Following infection, PrP^Sc circulates through the bloodstream and reaches secondary lymphoid tissues such as the lymph nodes, tonsils, Peyer’s patches, and spleen. At these sites, PrP^Sc utilizes immune cells to replicate and accumulate (46, 50). Once sufficient replication and accumulation occur, prion diseases are triggered subsequent to CNS infection, initiating the progression of pathological manifestations. Therefore, PrP^Sc accumulation in the secondary lymphoid tissues is crucial for movement of PrP^Sc into the CNS.

FDCs found in the B cell area of germinal centers (GCs) in secondary lymphoid tissues are critical for capturing naïve antigens using FcγRIIB and complement receptors and presenting them to GC B cells (51-53). In response to these antigens, B cells receive help from follicular helper T (Tfh) cells, which leads to their activation and subsequent differentiation into plasma cells (54-56). Due to high PrP^C expression levels, FDCs accumulate considerable amount of PrP^Sc upon infection. Consequently, FDCs are pivotal in the onset of prion diseases, and the spleen serves as the primary site for PrP^Sc replication mediated by FDCs (57-60). In the absence of FDCs, the lack of a site for accumulation of PrP^Sc prevents its accumulation. Consequently, neuroinvasion does not occur in the absence of PrP^Sc accumulation (60-62).

Our study showed that ME7-infected mice exhibited increased FDC networks and Tfh cell responses, which persisted throughout the progression of prion disease (42). Despite a decrease in CD4 T cells in the white pulp, there was an increase in CD4 T cells within GCs, accompanied by higher expression levels of Tfh-related genes, such as Bcl6, Il21, Cxcr5, Icos, and Pdcd1. Moreover, the ME7-infected spleens showed an elevated number of CD4 memory T cells. These results suggest that although ME7 infection led to an impaired structure in the splenic white pulp, there was an expansion of CD4 memory T cells and prolonged Tfh cell responses necessary to support the replication and accumulation of PrP^Sc within GCs.

ROLE OF PrP^C IN INFLAMMATION

Several studies have investigated the effects of PrP^C on inflammatory responses, given its substantial expression in immune cells and its ability to influence immune cell activation and recruitment. These studies revealed that PrP^C protects organs from inflammatory responses through its immunomodulatory function (63-67). High expression of PrP^C in immune-privileged organs, including the brain, placenta, eyes, testes, and uterus, serves as a protective mechanism against inflammation-induced damage to these organs (63). Inflammation studies using Prnp-knockout models have been conducted to investigate the protective role of PrP^C against inflammation (68-71). Tsutsui et al. demonstrated that Prnp^0/0 mice immunized with myelin oligodendrocyte glycoprotein peptide to induce experimental autoimmune encephalomyelitis exhibited a more aggressive disease onset characterized by higher levels of leukocytic infiltrates and increased expression of pro-inflammatory cytokine genes in the brain and spinal cord suggesting the protective role of PrP^C against neuroinflammation (68). Petit et al. investigated the severity of inflammatory bowel disease induced by dextran sodium sulfate and found that mice lacking PrP^C exhibited more severe symptoms than wild-type mice (69). Furthermore, when PrP^C was knocked down in human enterocytes, there was a decrease in cell-cell junctions. The weakening of the intestinal barrier, consisting of tight cell-cell junctions, can lead to vulnerability to external invasion and infection, potentially resulting in inflammatory bowel diseases such as Crohn’s disease and ulcerative colitis. Consistent with these findings, patients with Crohn’s disease or ulcerative colitis showed decreased levels of PrP^C at cell-cell junctions in the colonic epithelia (69). Taken together, these results revealed that in the absence of PrP^C, there was an increase in inflammatory responses and a greater extent of associated damage.

In another study, goats lacking PrP^C exhibited prolonged symptoms in response to lipopolysaccharide-induced (LPS)-induced systemic inflammation (70). In another study using LPS, Liu et al. showed that upon LPS injection, wild-type mice exhibited elevated levels of pro-inflammatory cytokines in the brain and spleen during the acute phase, whereas Prnp^0/0 mice displayed lower cytokine levels (71). Additionally, Prnp^0/0 mice showed higher mortality rates in response to LPS-induced septic shock. These results suggest that PrP^C plays a crucial role in protecting against LPS injection by modulating the inflammatory response.

ROLE OF PrP^C IN CANCER DEVELOPMENT AND METASTASIS

Several studies have demonstrated that PrP^C stimulates cancer cell proliferation through various mechanisms. Elevated levels of PrP^C expression have been linked to unfavorable prognoses and have been observed in various human cancers such as gastric carcinoma (72), renal adenocarcinoma (73), colorectal cancer (74, 75), breast cancer (76, 77), and melanoma (78). In gastric cancer cells, PrP^C overexpression activates the phosphatidylinositide 3-kinase pathway and upregulation of cyclin D1, promoting the G1/S phase transition and consequently facilitating cell proliferation (79). Another study reported that PrP^C influences the G1/S phase transition in several renal adenocarcinoma cell lines (80). Consistent with these findings, in the absence of PrP^C, the expression of cyclins and cyclin-dependent kinases is suppressed, leading to the inhibition of cell proliferation in colon cancer (81).

Gil et al. demonstrated that PrP^C contributes to the invasion and migration of breast cancer cells by regulating matrix metalloprotease-9 (MMP-9) (82). They showed that overexpression of PrP^C in the breast cancer cell line MCF-7 leads to an increase in MMP-9 expression by enhancing the association of NF-κB with the promoter region of the MMP-9 gene and activating the ERK signaling pathway. Conversely, when PrP^C is silenced using siRNA, a notable decrease in ERK activation and MMP-9 expression is observed, leading to the suppression of cell migration and invasion (82).

Reportedly, PrP^C plays a role in both cancer development and metastasis, the process by which cancer cells spread from the primary tumor to other parts of the body (72, 83, 84). Metastatic gastric cancers exhibit high PrP^C expression, which plays a substantial role in enhancing the adhesive, invasive, and metastatic capacities of gastric cancer cell lines (72). According to Pan et al., the N-terminal region of PrP^C can activate the MEK/ERK pathway, ultimately leading to transactivation of MMP11. This activation enhances the invasive and metastatic properties of gastric cancer cells, indicating their potential role in promoting metastasis (72). Additionally, the overexpression of MMP11 is frequently associated with a more aggressive tumor phenotype and resistance to apoptosis (83). Wang et al. showed that PrP^C is specifically expressed at the invasive front of colorectal cancers (CRCs), promoting tumor invasion through the acquisition of characteristics associated with epithelial-mesenchymal transition (84). Additionally, they showed that knockdown of PrP^C in an orthotopic xenograft model significantly reduced the number of distant metastases, supporting the significant role of PrP^C in the regulation of CRC progression and metastasis.

ROLE OF REGULATORY T CELLS IN CANCER AND THEIR RELATIONSHIP WITH PrP^C

Research on the role of PrP^C in cancer invasion and metastasis has revealed its association with regulatory T cells (37, 78). Regulatory T cells possess immunosuppressive functions and exhibit elevated activity in numerous types of cancers (85, 86). An increased number of tumor-infiltrating regulatory T cells have been identified, and their increased activity are observed in various human cancers, including liver, lung, breast, gastrointestinal tract, pancreas, and ovarian cancers (85-87). Our recent study showed that when B16F10 melanoma cells were injected into Prnp^0/0 and PrP-overexpressing (Tga20) mice to induce lung metastasis, Tga20 mice reached the terminal stage much faster than Prnp^0/0 mice as lung metastasis occurred (78). This effect was likely associated with an increased number of regulatory T cells. The expression of transforming growth factor-beta (TGF-β) and programmed death ligand-1 (PD-L1), which play important roles in the differentiation and function of regulatory T cells, was upregulated in Tga20 mice compared with Prnp^0/0 mice. These results suggest that during the invasion and metastasis of cancer cells, PrP^C may contribute to the development and activation of regulatory T cells by increasing the expression of TGF-β and PD-L1, which enhances their functionality.

CONCLUSION

In almost all cells, PrP^C is expressed, participating in diverse cellular processes and also found in immune cells, suggesting potential roles in the immune system. Based on the research findings to date, it has been revealed that PrP^C has a protective function in inflammatory responses triggered by infections, but contributes to the development of cancer through various mechanisms, including regulation of cell proliferation, enhancement of adhesive, invasive, and metastatic capacities, and boosting regulatory T cell activation. These findings suggest that PrP^C plays a crucial role in the immune system and has substantial implications in the pathogenesis of conditions like inflammation and cancer. However, since much of this research is based on results of phenomena observed through knockout systems, further investigations are required to unveil the specific roles of PrP^C in individual immune cells, elucidate molecular mechanisms, and understand the interactions through which PrP^C influences the immune system. Future studies exploring the precise functions of PrP^C in the immune system will enable the regulation of the immune system using PrP^C, which can be applied for the treatment of various diseases.

Funding Statement

ACKNOWLEDGEMENTS This work was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education, Science, and Technology (NRF- 2016R1D1A1B01011371).