Tumour necrosis factor receptor (TNFR)-associated factor (TRAF) is a critical linker molecule in the tumor necrosis factor superfamily (TNFSF) and Toll-like/interleukin-1 receptor (TLR/ILR) superfamily that plays a regulatory role in cell proliferation, differentiation, apoptosis and survival, and immune response. TRAF family members all have TRAF homologous structural domains at the carboxyl terminus, zinc-finger of the amino terminus, and ring finger structural domains. TRAF is a cytoplasmic bridging protein class that binds to other TRAF proteins after connecting to the receptor’s cytoplasmic domain. Six classical members (TRAF1-TRAF6) and one non-classical member (TRAF7) are known in mammals. TRAF2-6 has ring and zinc-finger motifs that are important for the downstream regulation of signaling (1). Structural features of TRAF proteins suggest that these proteins function as cytoplasmic junctions that promote intracellular signaling through their ability to bind to receptors and enhance the recruitment of proteins to signaling complexes (2).

In terms of structure, the TRAF-N region of the TRAF family is typically composed of more than ten 7-peptide repeats. In contrast, this region of TRAF4 contains only three 7-peptide repeats and the relatively short coiled helix structural domain of TRAF4 results in weak binding of the TRAF4 protein to other TRAF family proteins. Although the TRAF domain of TRAF4 has a typical TRAF domain fold, its high-resolution structure reveals similarities and differences between this structure and other TRAF family members, and these structural differences lead to functional variability (3). TRAF4 is the only TRAF family member with three CART structural domains, for which TRAF4 was once named CART1. Only TRAF4 in the TRAF family contains a unique N-terminal RING-finger and a nuclear localization signal (NLS), and TRAF4 may belong to a subfamily of TRAFs that function in the nucleus (4).

In terms of origin, the TRAF1, 2, 3 and 5 genes arose from the duplication of recent independent genes and had a common ancestor gene. Correlative evolutionary analyses show that TRAF1, 2, 3 and 5 emerged during vertebrate evolution. The TNFR family was formed at this same stage, suggesting that the functions of these four TRAF family proteins are related to those of the TNFR family proteins. Furthermore, TRAF1, 2, 3, 5 and 6 have been shown to interact directly or indirectly with the TNFR superfamily (5, 6). TRAF4 emerged earlier in evolution, homologous analogues of TRAF4 have been identified in lower coelenterates, and TRAF4 homologues play similar roles during embryonic development in both invertebrates and vertebrates, leading to the inference that TRAF4 may be one of the older members of the TRAF family and that it exerts a function that is TNFR non-dependent (7).

In terms of function, unlike other members of the TRAF protein family, TRAF4 migrates in the cell membrane, cell plasma and nucleus mainly through indirect action or formation of complexes, thus participating in several signalling pathways such as NF-κB and JNK to exert physiopathological regulatory functions. However, TRAF4 is less capable of activating signalling pathways alone and must be combined with other interacting proteins before it can effectively activate signalling pathways (8, 9).

Tumour necrosis factor (TNF) receptor-associated factor 4 (TRAF4) is an E3 ubiquitin ligase and a member of the tumor necrosis factor receptor-associated factor (TRAF) family, which is a crucial molecule in individual development. TRAF4 contains a nuclear localization signal (NLS), a unique TRAF family (4). The TRAF4 amino-terminal ring finger (RING) domain has E3 ubiquitin ligase activity, which promotes the ubiquitination of related proteins, regulates downstream signaling (10), and mediates activation of TAK1 and AKT1 target proteins and ubiquitination of K63 linkages (11). TRAF4 is involved in the transduction of multiple signaling pathways, and most of the biological effects of TRAF4 accompany the activation of signaling pathways activation ( Table 1 ).

TRAF4 is required during embryogenesis and is mainly involved in neurogenesis, cell polarity, cell proliferation, immunity, inflammation-related functions, apoptosis, oxidation, osteogenesis metabolism, and repair (12) ( Figure 1 ).

In neurogenesis, in the absence of TRAF4, increased phosphorylation of Rock2 (a downstream target of RhoA) leads to activation of the NgR/p75NTR/RhoA signaling pathway induces actin cytoskeletal rearrangement and facilitates axonal regeneration inhibition and neuronal apoptosis (13). TRAF4 is identified as a novel target of Smurf1 that polyubiquitinates TRAF4 to trigger its proteasomal destruction. TRAF4 enhances bone morphogenetic proteins BMP and Nodal signaling. TRAF4 functions as a novel Smurf1 regulatory mediator of BMP and Nodal signaling, essential for neural crest development and neural plate morphogenesis (14).

In cell polarity, TRAF4 is highly mobile and shuttles between the tight cell junction (TJ) and the cytoplasm. Intracellular TRAF4 enhances ERK1/2 phosphorylation in proliferating HeLa cells, an epithelial cell line that lacks TJ. TRAF4 is involved in an emerging TJ-dependent signaling pathway that affects cell polarity by regulating cell proliferation/differentiation homeostasis and subsequent epithelial homeostasis (15). TRAF4 contributes to TGF-β-induced epithelial-mesenchymal transition (EMT), metastasis, and p53 instability (16).

In cell proliferation, activation of the epidermal growth factor receptor (EGFR) induces changes in cellular functions such as proliferation, migration, and differentiation. TRAF4 binding induces conformational rearrangements in the JM region to promote EGFR dimerization, and TRAF4 overexpression promotes EGF-induced EGFR autophosphorylation and downstream signaling. In contrast, deletion of TRAF4 binding sites and specific point mutations significantly attenuated EGFR activation and EGF-driven cell proliferation (17).

In immunity, TRAF4 is involved in immune function by promoting immune cell migration (18). TRAF4 enhances glucocorticoid-induced TNFR (GITR), a receptor expressed on T cells, B cells, and macrophages, thereby triggering activation of NF-kappaB. TRAF4-mediated activation of NF-kappaB downstream of GITR depends on the TRAF binding site in the previously mapped receptor cytoplasmic domain and is inhibited by cytoplasmic protein A20. TRAF4 is thought to inhibit the suppressive function of regulatory T cells (Treg cells) and promote T cell activation. TRAF4 elaborates on GITR signaling and has been shown to regulate the suppressive function of Treg cells (19). TRAF4 binding motifs are involved in the termination of innate immune signaling responses (20).

In terms of inflammation-related functions, miR-4443 overexpression inhibited the TRAF4/Iκα/NF-κB signaling pathway to activate anti-inflammatory cytokine expression in monocytes. miR-4443 increased expression-induced monocyte dysfunction by targeting TRAF4, which may be a key mediator of SIDS (21). MiR-4443 directly inhibited TRAF 4 expression, increasing CD4+ T cell cytokine secretion and proliferation via the NF-κB pathway. Increased miR-4443 expression induces CD4+ T cell dysfunction by targeting TRAF4, leading to GD (22). TRAF4 is a critical negative regulator of NOD2 signaling. TRAF4 may be involved in regulating and coordinating the immune response to the intracellular pattern recognition receptor NOD2, which negatively regulates NOD2 through two consecutive glutamate residues of NOD2, leading to NF-κB pathway activation. TRAF4 inhibits NOD2-induced NF-κB activation and binds directly to NOD2 to inhibit NOD2-induced bacterial killing. TRAF4 phosphorylation and subsequent inhibition of NOD2 signaling require binding to the Crohn’s disease susceptibility protein NOD2 (23).

TRAF4 deficiency leads to tracheal malformations, resulting in altered airflow and increased lung inflammation (24). TRAF4 recruits the E3-ligase SMURF2 to degrade the IL-25R inhibitory molecule DAZAP2. TRAF4-SMURF2-mediated degradation of DAZAP2 is a crucial component of ACT1 recruitment to IL-25R to initiate IL-25 signaling and a critical step in airway inflammation (25). Increased TRAF4 expression in ankylosing spondylitis mesenchymal stem cells (MSCs) may impair lipopolysaccharide (LPS)-induced apoptotic autophagy by inhibiting Beclin-1 phosphorylation (26).

In terms of apoptosis, TRAF4 interacts with the dimeric neurotrophic factor receptor p75(NTR), and TRAF4 overexpression reduces its ability to dimerize to prevent the induction of apoptosis mediated by monomeric p75(NTR). TRAF4 also inhibits the NF activation response (27). TRAF4 is localized in the cytoplasm and appears to be retained in the cytoplasm after DNA damage Overexpression of TRAF4 induces apoptosis and inhibits colony formation (28).

In terms of oxidation, IL-17 and TNF-α induce upregulation of STEAP4 largely dependent on changes in TRAF4. TRAF4 is resistant to copper-mediated oxidative damage. The IL-17/TNF-α-TRAF4-STEAP4 axis can be used to regulate copper homeostasis (29). In TNF-α-stimulated endothelial cells (ECs), the binding of copper-dependent antioxidant-1 (Atox1) to TRAF4 leads to Atox1 nuclear translocation and induces a reactive oxygen species (ROS)-dependent inflammatory response (30). TRAF4 is directly associated with focal contact scaffolding (Hic-5), and knockdown of the protein, disruption of the complex, or oxidant scavenging prevents cell migration. Active mutants of TRAF4 activate Rho GTPases and p21-activated kinase 1 (PAK1) downstream of NADPH oxidase and oxidatively modify focal contact phosphatase (PTP-PEST). TRAF4 initiates robust membrane puckering via Rac1, PAK1, and oxidase, whereas knockdown of PTP-PEST increases membrane puckering while being unrelated to oxidase activation unrelated. TRAF4 specifies the molecular address in the focal complex targeting oxidative modifications during cell migration (31).

In terms of osteogenic metabolism, TRAF4 negatively regulates adipogenesis in MSCs through activation of PKM2 kinase activity, and downregulation of TRAF4 during adipogenesis is regulated by ALKBH5-mediated N6-methyladenosine RNA demethylation. This may serve as a checkpoint for fine-tuning the balance of adipose-osteogenic differentiation and suggests that TRAF4 may be a novel target for clinical MSCs and may also elucidate potential mechanisms of bone metabolic diseases (32). TRAF4 positively regulates osteogenesis in MSCs both in vitro and in vivo. TRAF4 acts as an E3 ubiquitin ligase that degrades Smurf2 to regulate the osteogenic differentiation of MSCs positively. TRAF4 plays a crucial role in bone formation to elucidate the pathogenesis of bone metabolism disorders and improve the practical clinical application of MSCs (33). TRAF4 acts as an E3-RING ubiquitin ligase, promotes the degradation of the adipocyte differentiation regulator PPARγ through the ubiquitin-proteasome pathway, thereby inhibiting adipogenesis.

In terms of repair, TRAF4 acts in a TNF receptor- and JNK-independent manner to fine-tune the assembly of adhesion junctions in invaginated mesangial cells (34). TRAF4 stimulates MAPK/ERK kinase 4 (MEKK4) activity by promoting MEKK4 oligomerization, and JNK activation can be facilitated by chemically inducing MEKK4 dimerization. MEKK4 is a TRAF4-regulated JNK pathway of MAPK kinase (9). TRAF4 inhibits the p53 pathway independent of its E3 ubiquitin ligase activity. TRAF4 interacts with the deubiquitinating enzyme USP10 and blocks p53 from entering USP10, resulting in p53 instability. TRAF4 proliferation regulation in skin repair is dependent on p53 (35). UPS is involved in skin repair by regulating the transforming growth factor (TGF)- β/Smad signaling pathway in the development and progression of hypertrophic scars. TRAF4 binding to the deubiquitinating enzyme USP10 induces p53 instability and promotes scar tissue-derived fibroblasts and skin repair (36).

Thus, by summarising, we can clearly elucidate that TRAF4 affects the physiological and pathological functions of tumor cells in appealing tumors by participating in signaling pathways. Through the NF-κB signaling pathway, TRAF4 promotes cancer cell proliferation in breast cancer, oesophageal cancer, osteosarcoma, and chronic lymphocytic leukemia. Through AKT or PI3K/AKT signaling pathway, TRAF4 promotes cancer cell migration and invasion in breast, liver, lung, endometrial, osteosarcoma, glioma, and ovarian cancers. Through the Wnt signaling pathway, TRAF4 promotes cancer cell invasion in colorectal cancer and glioma. In addition, through TNF-α signaling, TRAF4 exerts anti-apoptotic effects on breast cancer and osteosarcoma. Through the β-catenin signaling pathway, TRAF4 increases resistance to chemotherapeutic agents in breast and colorectal cancers. Through the TGF-β signaling pathway, TRAF4 promotes cancer cell migration and invasion in breast and prostate cancers. In breast cancer, TRAF4 promotes cancer cell migration and invasion through the Rac1 signaling pathway. In prostate cancer, TRAF4 promotes cancer cell proliferation through the RTK signaling pathway. In glioma, TRAF4 promotes cancer cell proliferation and invasion via the Hippo signaling pathway.

TRAF4 plays an essential role in physiological and pathological processes, which involves signalling pathways such as JNK, PI3K/AKT, MAPK/ERK, Wnt/β-catenin, TGF-β/Samd, NF-κB, Hippo and others. Exploring the interconnections between signalling pathways with a focus on TRAF4 will help understand the role of TRAF4 as an interacting protein and provide a reference for the use of TRAF4 in disease ( Figure 2 ).

TRAF4 promotes JNK activation by participating in the MAPK/ERK signaling pathway (9). TRAF4 binding to IRS-1 promotes its ubiquitination and phosphorylation, leading to activation of MAPK and PI3K/AKT signaling pathways. TRAF4 overexpression enhances IRS-1 tyrosine phosphorylation to encourage activation of ERK effector proteins (44). Increasing TRAF4 can promote NGF-stimulated activation of the p38 MAPK pathway downstream of TrkA and the expression of invasion-associated genes.

EMT induces TRAF4 to promote cancer cell migration and invasion by activating the Wnt/β catenin signaling pathway, and EMT induces p53 instability (16, 62). DR6 binding to TRAF4 enhances cancer cell mobility by enhancing MMP2 and MMP9 expression and participating in the PI3K/AKT pathway (82). TRAF4 involvement in the TGF-β/Samd signaling pathway can regulate MMP2, and MMP9 TRAF4 involvement in the TGF-β/Samd signaling pathway regulates the expression of MMP2 and MMP9 and induces P53 instability and tumorigenesis (16). Elevated TRAF4 expression regulates the TGF-β pathway and mediates an increase in phosphorylated SMAD2 and phosphorylated TAK1, triggering poor prognosis associated (11). In addition, TAK1 is involved in the NF-κB and JNK pathways and the p38 MAPK pathway (88).

Reducing TRAF4 can inhibit cancer cell proliferation, invasion and metastasis by downregulating the AKT signaling pathway, inhibiting the NF-κB pathway and engaging in RSK4 interactions (43). MiR-29a is an important tumor suppressor that promotes apoptosis and inhibits cell proliferation, migration and invasion by inhibiting the TRAF4/AKT/MDM2 pathway in a p53-dependent manner (77, 78). BCR promotes TRAF4 upregulation and activates the CD40-induced NF-κB signaling pathway through MYC inhibition of miR-29a expression (84). MiR-29a inhibits cell proliferation by blocking the phosphorylation of AKT and GSK-3β and cyclin D1 expression. High expression of TRAF4 significantly increased cyclinD1 and c-myc protein expression levels in cancer cells and activated the Wnt/β-catenin signaling pathway (75). DLEU1 can act as a competitive endogenous RNA that promotes elevated TRAF4 expression levels, promotes cancer cell proliferation through the Hippo signaling pathway and Wnt/β-catenin signaling pathway and regulate cell migration (79).

TRAF4 produces cell-specific and diverse biological effects in the regulation of cell differentiation, polarity, proliferation, and apoptosis, affecting embryonic development, rule of reactive oxygen species production, and mediating tumor formation and evolution. TRAF4 significantly enhances cancer development and progression in various malignancies, and its effects involve multiple signaling pathways such as AKT, NF-κB, Wnt, TGF-βand TNF-α. TRAF4 is a potential molecular target for cancer prevention and treatment, and the regulatory mechanism of TRAF4 needs to be further investigated. The study of TRAF4 can identify new targets suitable for tumor therapy and lay the foundation for developing new drugs and tumor drug resistance research.

XR wrote the review. RZ, RL, HZ, ZW, and CW discussed the review. All authors proofread the review. All authors contributed to the article and approved the submitted version.

This work was supported by the National Natural Science Foundation of China (82070175), Natural Science Foundation of Hunan Province (2021JJ30937), Scientific program of Health Commission of Hunan Province (2022030442723), Changsha Municipal Natural Science Foundation (kq2014234).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.