[Immune Regulation by TNF Receptor-associated Factor 5].

The tumor necrosis factor receptor (TNFR)-associated factor (TRAF) family of molecules are intracellular adaptors that regulate cellular signaling through members of the TNFR and Toll-like receptor superfamily. Mammals have seven TRAF molecules numbered sequentially from TRAF1 to TRAF7. Although TRAF5 was identified as a potential regulator of TNFR superfamily members, the in vivo function of TRAF5 has not yet been fully elucidated. We identified an unconventional role of TRAF5 in interleukin-6 (IL-6) receptor signaling involving CD4+ T cells. Moreover, TRAF5 binds to the signal-transducing glycoprotein 130 (gp130) receptor for IL-6 and inhibits the activity of the janus kinase (JAK)-signal transducer and activator of transcription (STAT) signaling pathway. In addition, Traf5-deficient CD4+ T cells exhibit significantly enhanced IL-6-driven differentiation of T helper 17 (Th17) cells, which exacerbates neuroinflammation in experimental autoimmune encephalomyelitis. Furthermore, TRAF5 demonstrates a similar activity to gp130 for IL-27, another cytokine of the IL-6 family. Additionally, Traf5-deficient CD4+ T cells display significantly increased IL-27-mediated differentiation of Th1 cells, which increases footpad swelling in delayed-type hypersensitivity response. Thus, TRAF5 functions as a negative regulator of gp130 in CD4+ T cells. This review aimed to explain how TRAF5 controls the differentiation of CD4+ T cells and discuss how the expression of TRAF5 in T cells and other cell types can influence the development and progression of autoimmune and inflammatory diseases.

Tumor necrosis factor receptor-associated factor 5 protects against intimal hyperplasia by regulation of macrophage polarization via directly targeting PPARγ.

OBJECTIVES: Intimal hyperplasia is a serious clinical problem associated with the failure of therapeutic methods in multiple atherosclerosis-related coronary heart diseases, which are initiated and aggravated by the polarization of infiltrating macrophages. The present study aimed to determine the effect and underlying mechanism by which tumor necrosis factor receptor-associated factor 5 (TRAF5) regulates macrophage polarization during intimal hyperplasia. METHODS: TRAF5 expression was detected in mouse carotid arteries subjected to wire injury. Bone marrow-derived macrophages, mouse peritoneal macrophages and human myeloid leukemia mononuclear cells were also used to test the expression of TRAF5 in vitro. Bone marrow-derived macrophages upon to LPS or IL-4 stimulation were performed to examine the effect of TRAF5 on macrophage polarization. TRAF5-knockout mice were used to evaluate the effect of TRAF5 on intimal hyperplasia. RESULTS: TRAF5 expression gradually decreased during neointima formation in carotid arteries in a time-dependent manner. In addition, the results showed that TRAF5 expression was reduced in classically polarized macrophages (M1) subjected to LPS stimulation but was increased in alternatively polarized macrophages (M2) in response to IL-4 administration, and these changes were demonstrated in three different types of macrophages. An in vitro loss-of-function study with TRAF5 knockdown plasmids or TRAF5-knockout mice revealed high expression of markers associated with M1 macrophages and reduced expression of genes related to M2 macrophages. Subsequently, we incubated vascular smooth muscle cells with conditioned medium of polarized macrophages in which TRAF5 expression had been downregulated or ablated, which promoted the proliferation, migration and dedifferentiation of VSMCs. Mechanistically, TRAF5 knockdown inhibited the activation of anti-inflammatory M2 macrophages by directly inhibiting PPARγ expression. More importantly, TRAF5-deficient mice showed significantly aggressive intimal hyperplasia. CONCLUSIONS: Collectively, this evidence reveals an important role of TRAF5 in the development of intimal hyperplasia through the regulation of macrophage polarization, which provides a promising target for arterial restenosis-related disease management.

METTL3-mediated m6 A methylation of TRAF5 inhibits lung adenocarcinoma cell metastasis via activation of the PI3K/AKT/NF-κB signaling pathway.

Tumor necrosis factor receptor-associated factor 5 (TRAF5) has been implicated in the pathogenesis of human malignancies. This work aimed to clarify the role of TRAF5 in lung adenocarcinoma (LUAD) progression. Herein, we uncovered that TRAF5 level was reduced in LUAD tissues. Low TRAF5 expression correlated with dismal prognosis in LUAD patients. Moreover, upregulated TRAF5 impeded cell viability, migration, and invasion, induced apoptosis in vitro, as well as impaired tumorigenicity in vivo. However, depletion of TRAF5 revealed opposing results. Moreover, TRAF5 was identified as the downstream target of methyltransferase-like 3 (METTL3)-elicited N6 -methyladenosine (m6 A) modification. METTL3 stabilized TRAF5 mRNA and positively modulated TRAF5 level. Further, TRAF5 depletion relieved the repressive phenotype caused by METTL3 addition. In addition, it was manifested that the METTL3/TRAF5 axis served as an inhibitor in LUAD through the PI3K/AKT/Nuclear Factor-Kappa B (NF-κB) signaling. Collectively, we propose that METTL3-mediated TRAF5 m6 A modification exerted as a vital tumor inhibitory function in LUAD development. The METTL3/TRAF5 axis may be a critical effector of LUAD progression.

TRAF5 regulates intestinal mucosal Th1/Th17 cell immune responses via Runx1 in colitis mice.

Inflammatory bowel disease (IBD) is a chronic gastrointestinal inflammatory disease associated with CD4+ Th1 and Th17 cell immune responses. Tumour necrosis factor-associated factor 5 (TRAF5) deficiency has been shown to aggravate DSS-induced colitis. However, the potential role of TRAF5 in regulating CD4+ T cell immune responses in the pathogenesis of IBD remains unclear. TRAF5-/- CD4+ CD45RBhigh T cells and WT CD4+ CD45RBhigh T cells were transferred to Rag2-/- mice via intravenous (i.v.) tail injection, respectively, to establish a chronic colitis model. Adeno-associated virus (AAV)-mediated gene knockout technique was used to knock out runt-associated transcription factor 1 (Runx1) expression in vivo. Specific cytokines of Th1 and Th17 cells were detected by quantitative RT-PCR, immunohistochemistry, ELISA, and flow cytometry. In T-cell transfer colitis mice, the Rag2-/- mice reconstituted with TRAF5-/- CD4+ CD45RBhigh T cells showed more severe intestinal inflammation than the WT control group, which was characterised by increased expression of INF-γ, TNF-α, IL-17a. Furthermore, we found that the INF-γ+ CD4+ , IL17a+ CD4+ , and INF-γ+ IL17a+ CD4+ T cells in the intestinal mucosa of Rag2-/- mice reconstituted with TRAF5-/- CD4+ CD45RBhigh T cells were significantly higher than those of the WT control group by flow cytometry. Mechanistically, knockout Runx1 inhibited the differentiation of TRAF5-/- CD4+ T cells into Th1 and Th17 cells in the intestinal mucosa of T-cell transfer colitis mice. TRAF5 regulates Th1 and Th17 cell differentiation and immune response through Runx1 to participate in the pathogenesis of colitis. Thus targeting TRAF5 in CD4+ T cells may be a novel treatment for IBD.

[Effect and mechanism of Bovis Calculus on ulcerative colitis by inhibiting IL-17/IL-17RA/Act1 signaling pathway].

This study aimed to elucidate the effect and underlying mechanism of Bovis Calculus in the treatment of ulcerative colitis(UC) through network pharmacological prediction and animal experimental verification. Databases such as BATMAN-TCM were used to mine the potential targets of Bovis Calculus against UC, and the pathway enrichment analysis was conducted. Seventy healthy C57BL/6J mice were randomly divided into a blank group, a model group, a solvent model(2% polysorbate 80) group, a salazosulfapyridine(SASP, 0.40 g·kg~(-1)) group, and high-, medium-, and low-dose Bovis Calculus Sativus(BCS, 0.20, 0.10, and 0.05 g·kg~(-1)) groups according to the body weight. The UC model was established in mice by drinking 3% dextran sulfate sodium(DSS) solution for 7 days. The mice in the groups with drug intervention received corresponding drugs for 3 days before modeling by gavage, and continued to take drugs for 7 days while modeling(continuous administration for 10 days). During the experiment, the body weight of mice was observed, and the disease activity index(DAI) score was recorded. After 7 days of modeling, the colon length was mea-sured, and the pathological changes in colon tissues were observed by hematoxylin-eosin(HE) staining. The levels of tumor necrosis factor-α(TNF-α), interleukin-1ß(IL-1ß), interleukin-6(IL-6), and interleukin-17(IL-17) in colon tissues of mice were detected by enzyme-linked immunosorbent assay(ELISA). The mRNA expression of IL-17, IL-17RA, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1ß, CXCL1, CXCL2, and CXCL10 was evaluated by real-time polymerase chain reaction(RT-PCR). The protein expression of IL-17, IL-17RA, Act1, p-p38 MAPK, and p-ERK1/2 was investigated by Western blot. The results of network pharmacological prediction showed that Bovis Calculus might play a therapeutic role through the IL-17 signaling pathway and the TNF signaling pathway. As revealed by the results of animal experiments, on the 10th day of drug administration, compared with the solvent model group, all the BCS groups showed significantly increased body weight, decreased DAI score, increased colon length, improved pathological damage of colon mucosa, and significantly inhibited expression of TNF-α,IL-6,IL-1ß, and IL-17 in colon tissues. The high-dose BCS(0.20 g·kg~(-1)) could significantly reduce the mRNA expression levels of IL-17, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1ß, CXCL1, and CXCL2 in colon tissues of UC model mice, tend to down-regulate mRNA expression levels of IL-17RA and CXCL10, significantly inhibit the protein expression of IL-17RA,Act1,and p-ERK1/2, and tend to decrease the protein expression of IL-17 and p-p38 MAPK. This study, for the first time from the whole-organ-tissue-molecular level, reveals that BCS may reduce the expression of pro-inflammatory cytokines and chemokines by inhibiting the IL-17/IL-17RA/Act1 signaling pathway, thereby improving the inflammatory injury of colon tissues in DSS-induced UC mice and exerting the effect of clearing heat and removing toxins.

PCSK9 inhibitor attenuates atherosclerosis by regulating SNHG16/EZH2/TRAF5-mediated VSMC proliferation, migration, and foam cell formation.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor has been demonstrated to exert a great cardioprotection in cardiometabolic impairments, including atherosclerosis. However, its underlying mechanism remains not fully understood. This study focuses on uncovering the actions of PCSK9 inhibitor on the connection between atherosclerosis and vascular smooth muscle cell (VSMC) behaviors. qRT-PCR was utilized to detect the expression of SNHG16. Proliferation and migration of VSMC were characterized by Cell Counting Kit-8 and wound healing assays. The intracellular lipids and foam cell formation were assessed by Oil Red O staining, fluorescence image, and cholesterol quantification kit. Atherosclerosis in vivo was evaluated by imaging the atherosclerotic lesions, hematoxylin-eosin staining, Oil Red O staining and Masson staining. The interaction between SNHG16 with EZH2 and histone H3 lysine 27 trimethylation (H3K27me3) were investigated by fluorescence in situ hybridization, RNA immunoprecipitation, and chromatin immunoprecipitation assays. A ApoE-/- mice model was used to validate the role of PCSK9 inhibitor and SNHG16 for atherosclerosis. The protective regulation of PCSK9 inhibitor was observed both in high-fat diet (HFD)-fed mice and oxidized low-density lipoprotein (ox-LDL)-treated VSMC, as manifested in the decreased the atherosclerotic lesions in vivo, as well as the weakened cell proliferation, migration, and formation of foam cells in vitro. SNHG16 was identified to be a downstream effector of PCSK9 inhibitor-mediated biological functions, of which knockdown also significantly ox-LDL-treated VSMC proliferation, migration, and foam cell formation abilities. SNHG16 epigenetically suppressed TRAF5 via recruiting EZH2. Silencing of TRAF5 abolished the protective effects of SNHG16 knockdown on the pathogenesis of atherosclerosis. Collectively, PCSK9 inhibitor attenuated atherosclerosis by regulating SNHG16/EZH2/TRAF5 axis to impair the proliferation, migration, and foam cell formation of VSMC.

Effect and mechanism of Bovis Calculus on ulcerative colitis by inhibiting IL-17/IL-17RA/Act1 signaling pathway / 中国中药杂志

This study aimed to elucidate the effect and underlying mechanism of Bovis Calculus in the treatment of ulcerative colitis(UC) through network pharmacological prediction and animal experimental verification. Databases such as BATMAN-TCM were used to mine the potential targets of Bovis Calculus against UC, and the pathway enrichment analysis was conducted. Seventy healthy C57BL/6J mice were randomly divided into a blank group, a model group, a solvent model(2% polysorbate 80) group, a salazosulfapyridine(SASP, 0.40 g·kg~(-1)) group, and high-, medium-, and low-dose Bovis Calculus Sativus(BCS, 0.20, 0.10, and 0.05 g·kg~(-1)) groups according to the body weight. The UC model was established in mice by drinking 3% dextran sulfate sodium(DSS) solution for 7 days. The mice in the groups with drug intervention received corresponding drugs for 3 days before modeling by gavage, and continued to take drugs for 7 days while modeling(continuous administration for 10 days). During the experiment, the body weight of mice was observed, and the disease activity index(DAI) score was recorded. After 7 days of modeling, the colon length was mea-sured, and the pathological changes in colon tissues were observed by hematoxylin-eosin(HE) staining. The levels of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), interleukin-6(IL-6), and interleukin-17(IL-17) in colon tissues of mice were detected by enzyme-linked immunosorbent assay(ELISA). The mRNA expression of IL-17, IL-17RA, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, CXCL2, and CXCL10 was evaluated by real-time polymerase chain reaction(RT-PCR). The protein expression of IL-17, IL-17RA, Act1, p-p38 MAPK, and p-ERK1/2 was investigated by Western blot. The results of network pharmacological prediction showed that Bovis Calculus might play a therapeutic role through the IL-17 signaling pathway and the TNF signaling pathway. As revealed by the results of animal experiments, on the 10th day of drug administration, compared with the solvent model group, all the BCS groups showed significantly increased body weight, decreased DAI score, increased colon length, improved pathological damage of colon mucosa, and significantly inhibited expression of TNF-α,IL-6,IL-1β, and IL-17 in colon tissues. The high-dose BCS(0.20 g·kg~(-1)) could significantly reduce the mRNA expression levels of IL-17, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, and CXCL2 in colon tissues of UC model mice, tend to down-regulate mRNA expression levels of IL-17RA and CXCL10, significantly inhibit the protein expression of IL-17RA,Act1,and p-ERK1/2, and tend to decrease the protein expression of IL-17 and p-p38 MAPK. This study, for the first time from the whole-organ-tissue-molecular level, reveals that BCS may reduce the expression of pro-inflammatory cytokines and chemokines by inhibiting the IL-17/IL-17RA/Act1 signaling pathway, thereby improving the inflammatory injury of colon tissues in DSS-induced UC mice and exerting the effect of clearing heat and removing toxins.

TNF Receptor-Associated Factor 5 Limits IL-27 Receptor Signaling in CD4+ T Lymphocytes.

TNF receptor-associated factor 5 (TRAF5) restrains early signaling activity of the IL-6 receptor in naive CD4+ T cells by interacting with the shared gp130 chain, although TRAF5 was initially discovered as a cytoplasmic adaptor protein to activate signaling mediated by TNF receptor family molecules. This leads to the question of whether TRAF5 limits signaling via the receptor for IL-27, which is composed of gp130 and WSX-1. The aim of this study is to clarify the role of TRAF5 in IL-27 receptor signaling and to understand the differential role of TRAF5 on cytokine receptor signaling. We found that Traf5 -/- CD4+ T cells displayed significantly higher levels of phosphorylated STAT1 and STAT-regulated genes Socs3 and Tbx21, as early as 1 h after IL-27 exposure when compared with Traf5 +/+ CD4+ T cells. Upon IL-27 and TCR signals, the Traf5 deficiency significantly increased the induction of IL-10 and promoted the proliferation of CD4+ T cells. Traf5 -/- mice injected with IL-27 displayed significantly enhanced delayed-type hypersensitivity responses, demonstrating that TRAF5 works as a negative regulator for IL-27 receptor signaling. In contrast, IL-2 and proliferation mediated by glucocorticoid-induced TNF receptor-related protein (GITR) and TCR signals were significantly decreased in Traf5 -/- CD4+ T cells, confirming that TRAF5 works as a positive regulator for cosignaling via GITR. Collectively, our results demonstrate that TRAF5 reciprocally controls signals mediated by the IL-27 receptor and GITR in CD4+ T cells and suggest that the regulatory activity of TRAF5 in gp130 is distinct from that in TNF receptor family molecules in a T cell.

Bioinformatics Analysis of miRNAs Targeting TRAF5 in DLBCL Involving in NF-κB Signaling Pathway and Affecting the Apoptosis and Signal Transduction.

Background: Diffuse large B-cell lymphoma (DLBCL) is an aggressive B-cell lymphoma with high heterogeneity. There is an unmet need to investigate valid indicators for the diagnosis and therapy of DLBCL. Methods: GEO database was utilized to screen for differentially expressed genes (DEGs) and differential miRNAs in DLBCL tissues. The Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) were applied to analyse DEGs. Then multiple databases were searched for related miRNAs within DLBCL, TNF receptor-associated factor 5 (TRAF5) and NF-kappa B (NF-κB) signaling pathways. The KOBAS database was used to assist in the screening of miRNAs of interest and construct the regulatory network of miRNA-mRNA. Finally, the expression level and diagnostic performance of miRNAs were analyzed with GEO datasets, and DEGs were identified from the GEPIA database. Results: DEGs were significantly concentrated in the NF-κB signaling pathway and cytokine-cytokine receptor interaction, and involved in the process of immune response and protein binding. MiR-15a-5p, miR-147a, miR-192-5p, miR-197-3p, miR-532-5p, and miR-650 were revealed to be targeting TRAF5 and participating in NF-κB signaling pathway and might impact the apoptosis and signal transduction of DLBCL. In the GEPIA database, TRAF5 was significantly overexpressed in DLBCL. The expression of miR-197-3p was upregulated within GEO datasets, while the rest of the miRNAs were downregulated in DLBCL. Conclusions: Subsets of miRNAs may participate in the NF-κB signaling pathway by co-targeting TRAF5 and could be prospective biomarkers exploring the pathogenesis of DLBCL.

Downregulation of long noncoding RNA SNHG7 protects against inflammation and apoptosis in Parkinson's disease model by targeting the miR-425-5p/TRAF5/NF-κB axis.

Accumulated evidence has manifested that long noncoding RNA (lncRNA) is involved in the progress of Parkinson's disease (PD). SNHG7, a novel lncRNA, has been found to be involved in tumorigenesis. However, SNHG7 expression and its functional effects on PD remain uncharted. Rotenone (Rot) was adopted to construct PD models in Sprague-Dawley (SD) rats and SH-SY5Y cells, respectively. The expression levels of caspase 3, tyrosine hydroxylase (TH), ionized calcium-binding adapter molecule 1 (Iba1) in SD rat striatum were measured via immunohistochemistry and western blot. Additionally, the expressions of inflammatory cytokines (interleukin 1ß [IL-1ß], IL-6, tumor necrosis factor α) and oxidative stress factors (malondialdehyde, superoxide dismutase, and glutathione peroxidase) in the brain tissues were examined using real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Moreover, the protein levels of tumor necrosis factor receptor-associated factor (TRAF5), I-κB, nuclear factor-κB (NF-κB), HO-1, Nrf2 were detected via western blot. Bioinformatics was applied to predict the targeting relationship between SNHG7, miR-425-5p, and TRAF5. Dual-luciferase activity assay and RNA immunoprecipitation assays were conducted to verify their interactions. In comparison to healthy donors, SNHG7 was found upregulated while miR-425-5p expression was downregulated in PD patients. Functional experiments confirmed that SNHG7 downregulation or miR-425-5p overexpression attenuated neuronal apoptosis in the Rot-mediated PD model, TH-positive cell loss, and microglial activation by mitigating inflammation and oxidative stress. Mechanistically, SNHG7 served as a competitive endogenous RNA by sponging miR-425-5p and promoted TRAF5 mediated inflammation and oxidative stress. Inhibition of SNHG7 ameliorated neuronal apoptosis in PD through relieving miR-425-5p/TRAF5/NF-κB signaling pathway modulated inflammation and oxidative stress, and similar results were observed in the Rot-mediated rat model of PD.

MicroRNA-29b-3p promotes 5-fluorouracil resistance via suppressing TRAF5-mediated necroptosis in human colorectal cancer.

Drug resistance in colorectal cancer is a great challenge in clinic. Elucidating the deep mechanism underlying drug resistance will bring much benefit to diagnosis, therapy and prognosis in patients with colorectal cancer. In this study, miR-29b-3p was shown to be involved in resistance to 5-fluorouracil (5-FU)-induced necroptosis of colorectal cancer. Further, miR-29b-3p was shown to target a regulatory subunit of necroptosis TRAF5. Rescue of TRAF5 could reverse the effect of miR-29b-3p on 5-FU-induced necroptosis, which was consistent with the role ofnecrostatin-1 (a specific necroptosis inhibitor). Then it was demonstrated that miR-29b-3p was positively correlated with chemo-resistance in colorectal cancer while TRAF5 negatively. In conclusion, it is deduced that miR-29b-3p/TRAF5 signaling axis plays critical role in drug resistance in chemotherapy for colorectal cancer patients by regulating necroptosis. The findings in this study provide us a new target for interfere therapy in colorectal cancer.

The Structure and Ubiquitin Binding Properties of TRAF RING Heterodimers.

Tumour necrosis factor (TNF) receptor associated factor (TRAF) family members share a common domain architecture, but play non-redundant physiological roles in cell signalling. At the N terminus, most TRAFs have a RING domain, followed by a series of Zinc finger (ZF) domains. The RING domain of TRAF6 dimerizes, and the RING homodimer together with the first ZF assembles ubiquitin chains that form a platform which facilitates activation of downstream kinases. The RING dimer interface is conserved amongst TRAF proteins, suggesting that functional heterodimers could be possible. Here we report the structure of the TRAF5-TRAF6 RING heterodimer, which accounts for the stability of the heterodimer as well as its ability to assemble ubiquitin chains. We also show that the RING domain of TRAF6 heterodimerizes with TRAF3 and TRAF2, and demonstrate that the linker helix and first ZF of TRAF2 can cooperate with TRAF6 to promote chain assembly. Collectively our results suggest that TRAF RING homo- and hetero-dimers have the potential to bridge interaction of nearby TRAF trimers and modulate TRAF-mediated signalling.

Down-regulated HDAC3 elevates microRNA-495-3p to restrain epithelial-mesenchymal transition and oncogenicity of melanoma cells via reducing TRAF5.

MicroRNAs (miRNAs) are emerging biomarkers in biological processes and the role of miR-495-3p has been identified in melanoma, while the detailed molecular mechanisms remain to be further explored. We aim to explore the effect of histone deacetylase 3 (HDAC3) and miR-495-3p on epithelial-mesenchymal transition (EMT) and oncogenicity of melanoma cells by regulating tumour necrosis factor receptor-associated factor 5 (TRAF5). Levels of HDAC3, miR-495-3p and TRAF5 in melanoma tissues and pigmented nevus tissues were determined, and the predictive roles of HDAC3 and miR-495-3p in prognosis of melanoma patients were measured. The melanoma cells were screened and transfected with relative oligonucleotides and plasmids, and the expression of HDAC3, miR-495-3p and TRAF5, and phenotypes of melanoma cells were gauged by a series of assays. The relations between HDAC3 and miR-495-3p, and between miR-495-3p and TRAF5 were confirmed. HDAC3 and TRAF5 were increased while miR-495-3p was decreased in melanoma cells and tissues, and the low expression of miR-495-3p as well as high expression of HDAC3 indicated a poor prognosis of melanoma patients. Inhibited HDAC3 elevated miR-495-3p to suppress EMT and oncogenicity of melanoma cells by reducing TRAF5. HDAC3 particularly bound to miR-495-3p and TRAF5 was the target gene of miR-495-3p. Our results revealed that down-regulated HDAC3 elevates miR-495-3p to suppress malignant phenotypes of melanoma cells by inhibiting TRAF5, thereby repressing EMT progression of melanoma cells. This study may provide novel targets for melanoma treatment.

TRAF5 protects against myocardial ischemia reperfusion injury via AKT signaling.

During the processes of myocardial ischemia reperfusion (I/R) injury, inflammation and apoptosis play an important role. I/R and its induced acute myocardial infarction (AMI) with high morbidity and mortality, and there is no effective treatment for it so far. TRAF5 has been shown to regulate inflammation and apoptosis in atherosclerosis, steatosis and melanoma cells, but its function in myocardial I/R injury is still unclear. This study demonstrates that the expression of TRAF5 is significant up-regulation in heart tissues of I/R injury mice and hypoxia/reoxygenation (H/R)-stimulated cardiomyocytes. TRAF5 knockout mice exhibites heavier heart damage, inflammatory response and cell death after myocardial I/R injury. Further, TRAF5 overexpression inhibites inflammation and apoptosis of H/R-stimulated cardiomyocytes. Mechanistically, we prove that TRAF5 promotes the activation of AKT. Overall, our study indicates that TRAF5 can regulate the processes of myocardial I/R injury. TRAF5 can be a new therapy target for myocardial I/R injury.

TRAF5 Deficiency Ameliorates the Severity of Dextran Sulfate Sodium Colitis by Decreasing TRAF2 Expression in Nonhematopoietic Cells.

TNFR-associated factor 5 (TRAF5) is a cytosolic adaptor protein and functions as an inflammatory regulator. However, the in vivo function of TRAF5 remains unclear, and how TRAF5 controls inflammatory responses in the intestine is not well understood. In this study, we found that intestinal epithelial cells from Traf5-/- mice expressed a significantly lower level of NF-κB-regulated proinflammatory genes, such as Tnf, Il6, and Cxcl1, as early as day 3 after dextran sulfate sodium (DSS) exposure when compared with wild-type mice. The intestinal barrier integrity of DSS-treated Traf5-/- mice remained intact at this early time point, and Traf5-/- mice showed decreased body weight loss and longer colon length at later time points. Surprisingly, the protein level of TRAF2, but not TRAF3, was reduced in colon tissues of Traf5-/- mice after DSS, indicating the requirement of TRAF5 for TRAF2 protein stability in the inflamed colon. Experiments with bone marrow chimeras confirmed that TRAF5 deficiency in nonhematopoietic cells caused the attenuated colitis. Our in vitro experiments demonstrated that proinflammatory cytokines significantly promoted the degradation of TRAF2 protein in Traf5-/- nonhematopoietic cells in a proteasome-dependent manner. Collectively, our data suggest a novel regulatory function of TRAF5 in supporting the proinflammatory function of TRAF2 in nonhematopoietic cells, which may be important for acute inflammatory responses in the intestine.

LINC00467 promotes cell proliferation and metastasis by binding with IGF2BP3 to enhance the mRNA stability of TRAF5 in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) has been recognized as a member of the most common human malignant tumors globally. According to multiple studies, long noncoding RNAs (lncRNAs) have been defined as vital regulators in tumor progression. Although previous studies have indicated that lncRNA long intergenic non-protein coding RNA 467 (LINC00467) exerts oncogenic effect in tumorigenesis and the development of cancers, the specific function that LINC00467 induces in HCC remains obscure. METHODS: LINC00467 expression was examined by a quantitative reverse transcriptase-polymerase chain reaction. CCK-8, EdU, transwell, western blotting and caspase-3 activity analyses were utilized to testify the role of LINC00467 in HCC. The interaction between IGF2BP3 and LINC00467 (or TRAF5) was investigated by luciferase reporter, RIP and RNA pull-down assays. RESULTS: LINC00467 upregulation in HCC tissues and cells was observed. LINC00467 silencing suppressed cell proliferation and metastasis, whereas it facilitated cell apoptosis in HCC. The gene for tumor necrosis factor receptor-associated factor 5 (TRAF5) was a neighboring gene of that for LINC00467 and its expression was positively modulated by LINC00467 in HCC. TRAF5 knockdown inhibited HCC progression. LINC00467 deficiency could decrease the mRNA stability of TRAF5 in HCC. Insulin-like growth factor-2 messenger RNA-binding protein 3 (IGF2BP3) could bind with LINC00467 and its depletion could lower TRAF5 mRNA stability in HCC. Final rescue assays further indicated that downregulation of IGF2BP3 or TRAF5 acted against LINC00467 upregulation-mediated function on HCC progression. CONCLUSIONS: LINC00467 promotes cell proliferation and metastasis by binding with IGF2BP3 to enhance the mRNA stability of TRAF5 in HCC.

miR-141-3p and TRAF5 Network Contributes to the Progression of T-Cell Acute Lymphoblastic Leukemia.

Numerous lines of evidence have shown that microRNAs (miRNAs) play a vital role in regulating the progression in many types of cancers, including T cell acute lymphoblastic leukemia (T-ALL). In this study, the potential underlying mechanism and functional role of miR-141-3p in T-ALL cells were determined. We found that the expression level of miR-141-3p was significantly downregulated, while that of tumor necrosis factor receptor-associated factor 5 (TRAF5) was strongly upregulated in tissues from patients with T-ALL compared with healthy controls. Subsequently, upregulation of miR-141-3p significantly repressed T-ALL cell proliferation and promoted cell apoptosis. Conversely, downregulation of miR-141-3p significantly inhibited cell apoptosis and enhanced T-ALL cell proliferation. We also verified that TRAF5 was the direct target of miR-141-3p in T-ALL cells. Additionally, TRAF5 overexpression significantly repressed cell apoptosis and increased T-ALL cell proliferation. In summary, miR-141-3p regulates T-ALL cell progression by directly targeting TRAF5, and may serve as a potential therapeutic target for T-ALL.

Overexpression of miRNA-410-3p protects hypoxia-induced cardiomyocyte injury via targeting TRAF5.

OBJECTIVE: This study aims to clarify the influence of microRNA-410-3p (miRNA-410-3p) on hypoxia-induced injury in cardiomyocytes. MATERIALS AND METHODS: MiRNA-410-3p level, apoptotic rate, and cell viability in AC16 cells undergoing normoxia or hypoxia preconditioning were assessed. The regulatory effects of miRNA-410-3p and TRAF5 on the proliferative and apoptotic abilities of AC16 cells were evaluated. The binding relationship between miRNA-410-3p and TRAF5 was verified by Dual-Luciferase Reporter Gene Assay. RESULTS: Hypoxia preconditioning triggered apoptosis and inhibited the viability in AC16 cells. MiRNA-410-3p was downregulated in cardiomyocytes under the hypoxic environment. The overexpression of miRNA-410-3p stimulated proliferation and inhibited apoptosis in hypoxia preconditioning AC16 cells. TRAF5 was proved to be the target of miRNA-410-3p. TRAF5 level was negatively regulated by miRNA-410-3p. The silence of TRAF5 could reverse viability and apoptosis changes in hypoxic AC16 cells overexpressing miRNA-410-3p. CONCLUSIONS: MiRNA-410-3p protects hypoxia-induced proliferation suppression and apoptosis stimulation in cardiomyocytes via targeting TRAF5.

TNF Receptor-Associated Factor 5 Limits Function of Plasmacytoid Dendritic Cells by Controlling IFN Regulatory Factor 5 Expression.

The physiological functions of TNF receptor-associated factor 5 (TRAF5) in the skin inflammation and wound healing process are not well characterized. We found that Traf5 -/- mice exhibited an accelerated skin wound healing as compared with wild-type counterparts. The augmented wound closure in Traf5 -/- mice was associated with a massive accumulation of plasmacytoid dendritic cells (pDCs) into skin wounds and an enhanced expression of genes related to wound repair at skin sites. In accordance with this result, adoptive transfer of Traf5 -/- pDCs, but not wild-type pDCs, into the injured skin area in wild-type recipient mice significantly promoted skin wound healing. The expression of skin-tropic chemokine receptor CXCR3 was significantly upregulated in Traf5-/- pDCs, and treatment with a CXCR3 inhibitor cancelled the promoted wound healing in Traf5-/- mice, suggesting a pivotal role of CXCR3 in pDC-dependent wound healing. Traf5 -/- pDCs displayed significantly higher expression of IFN regulatory factor 5 (IRF5), which correlated with greater induction of proinflammatory cytokine genes and CXCR3 protein after stimulation with TLR ligands. Consistently, transduction of exogeneous TRAF5 in Traf5-/- pDCs normalized the levels of abnormally elevated proinflammatory molecules, including IRF5 and CXCR3. Furthermore, knockdown of IRF5 also rescued the abnormal phenotypes of Traf5-/- pDCs. Therefore, the higher expression and induction of IRF5 in Traf5-/- pDCs causes proinflammatory and skin-tropic characteristics of the pDCs, which may accelerate skin wound healing responses. Collectively, our results uncover a novel role of TRAF5 in skin wound healing that is mediated by IRF5-dependent function of pDCs.

Tumor Necrosis Factor Receptor-Associated Factor 5 Promotes Arterial Neointima Formation through Smooth Muscle Cell Proliferation.

Tumor necrosis factor (TNF) receptor-associated factors (TRAFs) are cytoplasmic adaptor proteins of the TNF/interleukin (IL)-1/Toll-like receptor superfamily. Ligands of this family such as TNFα, CD40L, and IL-1ß promote chronic inflammatory processes such as atherosclerosis and restenosis, the latter being a common adverse reaction after vascular interventions. We previously reported overexpression of TRAF5 in murine and human atheromata and TRAF5-dependent proinflammatory functions in vitro. However, the role of TRAF5 in restenosis remains unsettled. To evaluate whether TRAF5 affects neointima formation, TRAF5-/-LDLR-/- and TRAF5+/+LDLR-/- mice consuming a high cholesterol diet (HCD) received wire-induced injury of the carotid artery. After 28 days, TRAF5-deficient mice showed a 45% decrease in neointimal area formation compared with TRAF5-compentent mice. Furthermore, neointimal vascular smooth muscle cells (vSMC) and macrophages decreased whereas collagen increased in TRAF5-deficient mice. Mechanistically, the latter expressed lower transcript levels of the matrix metalloproteinases 2 and 9, both instrumental in extracellular matrix degradation and vSMC mobilization. Additionally, TRAF5-specific siRNA interference rendered murine vSMC less proliferative upon CD40L stimulation. In accordance with these findings, fewer vSMC isolated from TRAF5-deficient aortas were in a proliferative state as assessed by Ki67 and cyclin B1 expression. In conclusion, TRAF5 deficiency mitigates neointima formation in mice, likely through a TRAF5-dependent decrease in vSMC proliferation.