Proteasome Subunit Beta Type 8 Research Articles

ZNF655 promotes the progression of hepatocellular carcinoma through PSMB8.

Hepatocellular carcinoma (HCC) is a type of liver cancer that is associated with high mortality rates. This study aims to investigate the role of ZNF655, a member of the zinc finger protein family, in the development of HCC. Immunohistochemical staining analysis was conducted to evaluate the expression of ZNF655 in HCC patient samples. Lentivirus-mediated ZNF655 knockdown was established in HCC cell lines (BEL-7402 and HCCLM3). The effects of ZNF655 on different aspects of HCC cell behavior such as proliferation, apoptosis, cycle, migration and tumor formation were examined. Downstream targets of ZNF655 in HCC were identified and verified through loss/gain-of-function experiments. Clinically, ZNF655 expression was elevated in HCC and increased with the severity of the disease. Functionally, inhibition of ZNF655 expression reduced the progression of HCC cells by decreasing proliferation, causing apoptosis, arresting cell cycle retention in G2, suppressing migration, and attenuating tumor formation in mice. Mechanistically, the proteasome subunit beta type-8 (PSMB8) was found to be co-expressed with ZNF655 in HCC, and PSMB8 knockdown weakened the promotion of ZNF655 overexpression on HCC. In summary, these findings suggest that ZNF655 promotes the progression of HCC through PSMB8, and inhibition of its expression may be a promising therapeutic target for HCC.

ONX-0914 Induces Apoptosis and Autophagy with p53 Regulation in Human Glioblastoma Cells.

Glioblastoma is believed to be one of the most aggressive brain tumors in the world. ONX-0914 (PR957) is a selective inhibitor of proteasome subunit beta type-8 (PSMB8). Previous studies have shown that inhibiting PSMB8 expression in glioblastoma reduces tumor progression. Therefore, this study aimed to determine whether ONX-0914 has antitumor effects on human glioblastoma. The results indicated that ONX-0914 treatment inhibited survival in LN229, GBM8401, and U87MG glioblastoma cells. Cell cycle analysis showed that ONX-0914 treatment caused cell cycle arrest at the G1 phase and apoptosis in glioblastoma cells. The protein expression of BCL-2 was reduced and PARP was cleaved after ONX-0914 treatment. Furthermore, the levels of p53 and phosphorylated p53 were increased by ONX-0914 treatment in glioblastoma cells. ONX-0914 also induced autophagy in glioblastoma cells. Furthermore, the p53 inhibitor pifithrin attenuated apoptosis but enhanced autophagy caused by ONX-0914. In an orthotopic mouse model, TMZ plus ONX-0914 reduced tumor progression better than the control or TMZ alone. These data suggest that ONX-0914 is a novel therapeutic drug for glioblastoma.

Comparative Transcriptomic Analysis Reveals the Regulated Expression Profiles in Oreochromis niloticus in Response to Coinfection of Streptococcus agalactiae and Streptococcus iniae.

Tilapia (Oreochromis sp.) is one of the important economical fishes in the world. Streptococcosis is commonly found in tilapia, causing severe and devastating effects in tilapia cultures. Streptococcus agalactiae and Streptococcus iniae are the predominant pathogens causing tilapia streptococcosis. To understand the molecular mechanisms underlying differential streptococcal infection patterns, Nile tilapias (Oreochromis niloticus) were infected by 1 × 107 CFU/mL S. agalactiae, 1 × 107 CFU/mL S. iniae, and 1 × 107 CFU/mL S. agalactiae and S. iniae (1:1), respectively, and transcriptome analysis was conducted to the intestine samples of Nile tilapia (Oreochromis niloticus) at 6, 12, 24 h, and 7 days post-infection. A total of 6,185 genes that differentially expressed among groups were identified. Eight differentially expressed genes (DEGs) including E3 ubiquitin-protein ligase TRIM39-like, C-X-C motif chemokine 10-like(CXCL 10), C-C motif chemokine 19-like, interleukin-1 beta-like, IgM heavy chain VH region, partial, IgG Fc-binding protein, proteasome subunit beta type-8 (PSMB8), and ATP synthase F(0) complex subunit B1, mitochondrial that involved in the immune system were selected, and their expression levels in the coinfection group were significantly higher than those in either of the single infection groups. These genes were associated with four different KEGG pathways. Additionally, the differential expression of eight DEGs was validated by using the RT-qPCR approach, and their immunological importance was discussed. The results provided insights into the responses of tilapia against S. agalactiae and S. iniae at the transcriptome level, promoting our better understanding of immune responses for aquatic animal against Streptococcus.

Proteasome subunit beta type-8 from sevenband grouper negatively regulates cytokine responses by interfering NF-κB signaling upon nervous necrosis viral infection

During viral infection, proper regulation of immune signaling is essential to ensure successful clearance of virus. Immunoproteasome is constitutively expressed and gets induced during viral infection by interferon signaling and contributes to regulate proinflammatory cytokine production and activation of the NF-κB pathway. In this study, we identified Hs-PSMB8, a member of the proteasome β-subunits (PSMB) family, as a negative regulator of NF-κB responses during NNV infection. The transient expression of Hs-PSMB8 delayed the appearance of cytopathic effect (CPE) and showed a higher viral load. The Hs-PSMB8 interacted with NNV which was confirmed using immunocolocalization and co-IP. Overexpression of Hs-PSMB8 diminished virus induced activation of the NF-κB promoters and downregulated the activation of IL-1β, TNFα, IL6, IL8, IFNγ expression upon NNV infection. Collectively, our results demonstrate that PSMB8 is an important regulator of NF-κB signaling during NNV infection in sevenband grouper.

Galectin-9 and PSMB8 overexpression predict unfavorable prognosis in patients with AML.

Acute myeloid leukemia (AML) is a deadly heterogeneous hematologic malignancy. Despite the well-characterized genetic characteristics and new promising targeted therapies for AML, the clinical outcome remains suboptimal. Galectin-9 (Gal-9) is a good potential target due to its immunosuppressive capacity in inflammatory processes. In our study, we firstly performed a wide range of integrated bioinformatical approach to assess the importance of Gal-9 by analyzing the expression, potential function and prognostic impact in AML. The results indicated that Gal-9 is overexpressed in AML cells, especially when relapse after hematopoietic stem cell transplantation (HSCT) and predicts poor prognosis. Co-expression analysis showed Gal-9 has a strong positive correlation with proteasome subunit beta type-8 (PSMB8), which was also highly expressed in AML with poor prognosis, implying a synergy in cell survival, cell signaling and the development of AML. In summary, we have confirmed the overexpression of Gal-9 and its partner PSMB8 in AML and validated their importance as prognostic factors. We propose that Gal-9 and PSMB8 could be a promising molecular target for treatment of AML and may provide more combined treatment options, especially in patients with relapse after HSCT.

PSMB8 inhibition decreases tumor angiogenesis in glioblastoma through vascular endothelial growth factor A reduction.

Glioblastoma, also known as glioblastoma multiforme (GBM), is a fast‐growing tumor and the most aggressive brain malignancy. Proteasome subunit beta type‐8 (PSMB8) is one of the 17 essential subunits for the complete assembly of the 20S proteasome complex. The aim of the present study was to evaluate the role of PSMB8 expression in GBM progression and angiogenesis. PSMB8 expression in glioblastoma LN229 and U87MG was knocked down by siRNA or inducible shRNA both in vitro and in vivo. After PSMB8 reduction, cell survival, migration, invasion, angiogenesis, and the related signaling cascades were evaluated. An orthotopic mouse tumor model was also provided to examine the angiogenesis within tumors. A GEO profile analysis indicated that high expression of PSMB8 mRNA in GBM patients was correlated with a low survival rate. In immunohistochemistry analysis, PSMB8 expression was higher in high‐grade than in low‐grade brain tumors. The proliferation, migration, and angiogenesis of human GBM cells were decreased by PSMB8 knockdown in vitro. Furthermore, phosphorylated focal adhesion kinase (p‐FAK), p‐paxillin, MMP2, MMP9, and cathepsin B were significantly reduced in LN229 cells. Integrin β1 and β3 were reduced in HUVEC after incubation with LN229‐conditioned medium. In an orthotopic mouse tumor model, inducible knockdown of PSMB8 reduced the expression of vascular endothelial growth factor (VEGF), VEGF receptor, and CD31 as well as the progression of human glioblastoma. In this article, we demonstrated the role of PSMB8 in glioblastoma progression, especially neovascularization in vitro and in vivo. These results may provide a target for the anti–angiogenic effect of PSMB8 in glioblastoma therapy in the future.

MicroRNA analysis of childhood atopic dermatitis reveals a role for miR-451a.

MicroRNAs (miRNAs), important regulators of gene expression, have been implicated in a variety of disorders. The expression pattern of miRNAs in paediatric atopic dermatitis (AD) has not been well studied. We sought to investigate miRNA expression profiles in different blood compartments of infants with AD. Small RNA and analysis with the HTG EdgeSeq system were performed to identify differentially expressed miRNAs in peripheral blood mononuclear cells (PBMCs) and plasma of infants with AD vs. age-matched healthy controls, with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) used for validation and measurement of miRNA targets. Logistic regression models with area under the receiving operating characteristic estimation was used to evaluate the diagnostic potential of chosen miRNAs for AD. RNA sequencing was performed to access miRNA expression profiles in paediatric AD. We identified 10 differentially expressed miRNAs in PBMCs and eight dysregulated miRNAs in plasma of infants with AD compared with controls. Upregulated miRNAs in PBMCs included miRNAs known to be involved in inflammation: miR-223-3p, miR-126-5p and miR-143-3p. Differential expression of only one miRNA, miR-451a, was observed in both PBMCs and plasma of children with AD. Dysregulation of three miRNAs (miR-451a, miR-143-3p and miR-223-3p) was validated in larger numbers of samples and miR-451a was identified as a predictive biomarker for the early diagnosis of the disease. Experimentally verified targets of miR-451a, interleukin 6 receptor (IL6R) and proteasome subunit beta type-8 (PSMB8), were increased in patients with AD, negatively correlated with miR-451a levels and upregulated following inhibition of miR-451a in PBMCs. In infants with AD, a distinct peripheral blood miRNA signature is seen, highlighting the systemic effects of the disease. miR-451a is uniquely expressed in different blood compartments of patients with AD and may serve as a promising novel biomarker for the early diagnosis of AD.

A novel mouse model of proteasome associated autoinflammatory syndromes

Abstract Proteasome associated autoinflammatory syndromes (PRAAS) represent a rare class of early-onset autoinflammatory diseases, including chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE syndrome). CANDLE is an inherited autosomal recessive disease associated with recurrent fevers, skin lesions, lipodystrophy and other comorbidities. Point mutations in the proteasome subunit beta type-8 (psmb8) gene, which encodes the beta-5i (ß5i) catalytic subunit of the 20S immunoproteasome, have been linked to CANDLE. The immunoproteasome is induced in cells in response to interferon (IFN) signaling. In cells lacking functional immunoproteasomes, oxidant-damaged proteins aggregate and likely create a feed-forward loop resulting in greater cytokine-mediated stress. We have developed a novel mouse model via targeted mutation within the psmb8 gene, which harbors a specific point mutation (224C>T) present in CANDLE patients. Our data shows that this mutation leads to marked differences in the expression of the immunoproteasome subunit ß5i within these mice. Thus, this mouse model is likely to exhibit changes in the immunoproteasome consistent with human disease upon interferon signaling. As mice with the targeted mutation do not directly yield a CANDLE-like phenotype at rest, it suggests that induction of disease-associated morbidity requires additional drivers. To test this, we are investigating interferons as initiators of disease pathogenesis. Future research will entail subjecting these mice to viral infection in order to identify if such triggers can initiate PRAAS-like symptoms in CANDLE patients. These mice may prove to an essential tool in understanding such disorders.

LncRNA HCP5 promotes LAML progression via PSMB8-mediated PI3K/AKT pathway activation.

Acute myeloid leukemia is an aggressive myeloid malignancy, characterized by rapid cellular proliferation and generally high mortality. Due to the lack of a complete understanding of AML, its clinical outcomes are still not satisfactory. In this study, we examined the function of the long non-coding RNA-HLA complex P5 (HCP5) on AML by analyzing the clinical samples, TCGA data, and by shRNA-mediated HCP5 deficiency in vitro. Our results showed that HCP5 is highly expressed in AML and is positive associated with poor prognosis, and HCP5 knockdown was significantly suppressing AML cell line proliferation and inducing G1/S arrest in vitro. In mechanism, the proteasome subunit beta type 8 (PSMB8) expression was dramatically inhibited in HCP5 knockdown cells while increased in HCP5 overexpression cells. PSMB8 was also highly expressed in AML and with poor prognosis. Furthermore, HCP5 regulates PI3K/AKT pathway activation depending on PSMB8. Our results showed a promoting function of HCP5 on AML and may provide a compelling biomarker and therapy target for AML.

MiR‐451a inhibits cancer growth, epithelial‐mesenchymal transition and induces apoptosis in papillary thyroid cancer by targeting PSMB8

Despite the increasing incidence of papillary thyroid cancer in the past decade, the molecular mechanism underlying its progression remains unknown. Several studies have reported down‐regulation of miR‐451a or circular miR‐451a in papillary thyroid cancer cell lines or patients. However, the underlying molecular mechanism remains unknown. In this study, we found that overexpression of miR‐451a could inhibit proliferation, epithelial‐mesenchymal transition and induce apoptosis in papillary thyroid cancer cells. Proteasome subunit beta type‐8 was predicted to be a direct target of miR‐451a and was validated with a luciferase reporter assay. Further functional assays showed that miR‐451a could inhibit thyroid cancer progression by targeting proteasome subunit beta type‐8.

Transcriptome profiling of antiviral immune and dietary fatty acid dependent responses of Atlantic salmon macrophage-like cells

BackgroundDue to the limited availability and high cost of fish oil in the face of increasing aquaculture production, there is a need to reduce usage of fish oil in aquafeeds without compromising farm fish health. Therefore, the present study was conducted to determine if different levels of vegetable and fish oils can alter antiviral responses of salmon macrophage-like cells (MLCs). Atlantic salmon (Salmo salar) were fed diets containing 7.4% (FO7) or 5.1% (FO5) fish oil. These diets were designed to be relatively low in EPA + DHA (i.e. FO7: 1.41% and FO5: 1%), but near the requirement level, and resulting in comparable growth. Vegetable oil (i.e. rapeseed oil) was used to balance fish oil in experimental diets. After a 16-week feeding trial, MLCs isolated from fish in these dietary groups were stimulated by a viral mimic (dsRNA: pIC) for 6 h (qPCR assay) and 24 h (microarray and qPCR assays).ResultsThe fatty acid composition of head kidney leukocytes varied between the two dietary groups (e.g. higher 20:5n-3 in the FO7 group). Following microarray assays using a 44K salmonid platform, Rank Products (RP) analysis showed 14 and 54 differentially expressed probes (DEP) (PFP < 0.05) between the two diets in control and pIC groups (FO5 vs. FO7), respectively. Nonetheless, Significance Analysis of Microarrays (SAM, FDR < 0.05) identified only one DEP between pIC groups of the two diets. Moreover, we identified a large number (i.e. 890 DEP in FO7 and 1128 DEP in FO5 overlapping between SAM and RP) of pIC-responsive transcripts, and several of them were involved in TLR−/RLR-dependent and cytokine-mediated pathways. The microarray results were validated as significantly differentially expressed by qPCR assays for 2 out of 9 diet-responsive transcripts and for all of the 35 selected pIC-responsive transcripts.ConclusionFatty acid-binding protein adipocyte (fabp4) and proteasome subunit beta type-8 (psmb8) were significantly up- and down-regulated, respectively, in the MLCs of fish fed the diet with a lower level of fish oil, suggesting that they are important diet-responsive, immune-related biomarkers for future studies. Although the different levels of dietary fish and vegetable oils involved in this study affected the expression of some transcripts, the immune-related pathways and functions activated by the antiviral response of salmon MLCs in both groups were comparable overall. Moreover, the qPCR revealed transcripts responding early to pIC (e.g. lgp2, map3k8, socs1, dusp5 and cflar) and time-responsive transcripts (e.g. scarb1-a, csf1r, traf5a, cd80 and ctsf) in salmon MLCs. The present study provides a comprehensive picture of the putative molecular pathways (e.g. RLR-, TLR-, MAPK- and IFN-associated pathways) activated by the antiviral response of salmon MLCs.

Evolution of dimorphisms of the proteasome subunit beta type 8 gene (PSMB8) in basal ray-finned fish

The proteasome subunit beta type 8 (PSMB8) gene encodes a catalytic subunit of immunoproteasome that plays a central role in the processing of antigenic peptides presented by major histocompatibility complex class I molecules. The A- and F-type alleles defined by the 31st amino acid residue determining cleaving specificity have been identified from ray-finned fish, amphibia, and reptiles. These two types show extremely long-term trans-species polymorphism in Polypteriformes, Cypriniformes, and Salmoniformes, suggesting the presence of very ancient lineages termed A and F. To elucidate the evolution of the PSMB8 dimorphism in basal ray-finned fish, we analyzed Pantodon buchholzi (Osteoglossiformes), seven species of Anguilliformes, and Hypomesus nipponensis (Osmeriformes). Both A and F lineage sequences were identified from P. buchholzi and H. nipponensis, confirming that these two lineages have been conserved by basal ray-finned fish. However, both the A- and F-type alleles found in Anguilliformes species belonged to the F lineage irrespective of their types. This apparently suggests that the A lineage was lost in the common ancestor of Anguilliformes, and recovery of the A type within the F lineage occurred in Anguilliformes. The apparent loss of the F lineage and recovery of the F type within the A lineage have already been reported from tetrapods and higher teleosts. However, this is the first report on the reverse situation and reveals the dynamic evolution of the PSMB8 dimorphism.

Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature syndrome: a report of a novel mutation and review of the literature

Dear Editor, Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) syndrome is a new autoinflammatory syndrome caused by mutations in the gene for proteasome subunit, beta type, 8 (PSMB8). We report a young adult with a novel homozygous PSMB8 mutation and marked skin disease.

THU0371 Increased expression of inflammatory cytokines and chemokines in an autoinflammatory disorder, nakajo-nishimura syndrome

BackgroundThe ubiquitin-proteasome pathway is responsible for selective degradation of short-lived, misfolded and toxic proteins. It plays an important rule in transcriptional regulation.Nakajo-Nishimura syndrome (NNS) (MIM 256040) is an autosomal recessive...

Long-Lived Dichotomous Lineages of the Proteasome Subunit Beta Type 8 (PSMB8) Gene Surviving More than 500 Million Years as Alleles or Paralogs

On an evolutionary time scale, polymorphic alleles are believed to have a short life, persisting at most tens of millions of years even under long-term balancing selection. Here, we report highly diverged trans-species dimorphism of the proteasome subunit beta type 8 (PSMB8) gene, which encodes a catalytic subunit of the immunoproteasome responsible for the generation of peptides presented by major histocompatibility complex (MHC) class I molecules, in lower teleosts including Cypriniformes (zebrafish and loach) and Salmoniformes (trout and salmon), whose last common ancestor dates to 300 Ma. Moreover, phylogenetic analyses indicated that these dimorphic alleles share lineages with two shark paralogous genes, suggesting that these two lineages have been maintained for more than 500 My either as alleles or as paralogs, and that conversion between alleles and paralogs has occurred at least once during vertebrate evolution. Two lineages termed PSMB8A and PSMB8F show an A(31)F substitution that would probably affect their cleaving specificity, and whereas the PSMB8A lineage has been retained by all analyzed jawed vertebrates, the PSMB8F lineage has been lost by most jawed vertebrates except for cartilaginous fish and basal teleosts. However, a possible functional equivalent of the PSMB8F lineage has been revived as alleles within the PSMB8A lineage at least twice during vertebrate evolution in the amphibian Xenopus and teleostean Oryzias species. Dynamic evolution of the PSMB8 polymorphism through long-term persistence, loss, and regaining of dimorphism and conversion between alleles and paralogs implies the presence of strong selective pressure for functional polymorphism of this gene.

Highly divergent dimorphic alleles of the proteasome subunit beta type-8 (PSMB8) gene of the bichir Polypterus senegalus: implication for evolution of the PSMB8 gene of jawed vertebrates

The proteasome subunit beta type-8 (PSMB8) gene encodes a catalytic subunit of the immunoproteasome, which is involved in the generation of peptides presented by MHC class I molecules. To date, highly diverged dichotomous alleles of PSMB8 have been reported in Oryzias species (actinopterygian teleosts) and Xenopus species (sarcopterygian amphibians). These dimorphic alleles share a similar substitution (A/V(31)F/Y) at the 31st position of the mature protein, which is most probably involved in formation of the S1 pocket. This substitution likely confers different cleavage specificities on the dimorphic PSMB8s. In addition, two paralogous PSMB8 genes possessing the A and F residues at the 31st position have been reported in sharks. Phylogenetic analysis indicated that the two types of PSMB8 of Oryzias, Xenopus, and sharks arose by independent evolutionary events. Here, we identified another pair of dimorphic alleles of PSMB8, which have the A and F residues at the 31st position of the mature protein, from bichir, Polypterus senegalus, a basal actinopterygian. The sequences of the mature proteins-encoding region of the dimorphic alleles of bichir PSMB8, the A and F types, showed only 72.7% and 77.5% identities at the nucleotide and the deduced amino acid levels, respectively. Their intronic sequences show almost no similarity, indicating that the dimorphic alleles of bichir PSMB8 have a very ancient origin. However, phylogenetic analysis showed that the dimorphisms of PSMB8 of bichir, Xenopus, and Oryzias arose by independent evolutionary events, suggesting the presence of a strong selective pressure for possessing the dimorphism.

Proteasome assembly defect due to a proteasome subunit beta type 8 (PSMB8) mutation causes the autoinflammatory disorder, Nakajo-Nishimura syndrome

Nakajo-Nishimura syndrome (NNS) is a disorder that segregates in an autosomal recessive fashion. Symptoms include periodic fever, skin rash, partial lipomuscular atrophy, and joint contracture. Here, we report a mutation in the human proteasome subunit beta type 8 gene (PSMB8) that encodes the immunoproteasome subunit β5i in patients with NNS. This G201V mutation disrupts the β-sheet structure, protrudes from the loop that interfaces with the β4 subunit, and is in close proximity to the catalytic threonine residue. The β5i mutant is not efficiently incorporated during immunoproteasome biogenesis, resulting in reduced proteasome activity and accumulation of ubiquitinated and oxidized proteins within cells expressing immunoproteasomes. As a result, the level of interleukin (IL)-6 and IFN-γ inducible protein (IP)-10 in patient sera is markedly increased. Nuclear phosphorylated p38 and the secretion of IL-6 are increased in patient cells both in vitro and in vivo, which may account for the inflammatory response and periodic fever observed in these patients. These results show that a mutation within a proteasome subunit is the direct cause of a human disease and suggest that decreased proteasome activity can cause inflammation.

PSMB8 Encoding the β5i Proteasome Subunit Is Mutated in Joint Contractures, Muscle Atrophy, Microcytic Anemia, and Panniculitis-Induced Lipodystrophy Syndrome

We performed homozygosity mapping in two recently reported pedigrees from Portugal and Mexico with an autosomal-recessive autoinflammatory syndrome characterized by joint contractures, muscle atrophy, microcytic anemia, and panniculitis-induced lipodystrophy (JMP). This revealed only one homozygous region spanning 2.4 Mb (5818 SNPs) on chromosome 6p21 shared by all three affected individuals from both families. We directly sequenced genes involved in immune response located in this critical region, excluding the HLA complex genes. We found a homozygous missense mutation c.224C>T (p.Thr75Met) in the proteasome subunit, beta-type, 8 (PSMB8) gene in affected patients from both pedigrees. The mutation segregated in an autosomal-recessive fashion and was not detected in 275 unrelated ethnically matched healthy subjects. PSMB8 encodes a catalytic subunit of the 20S immunoproteasomes called β5i. Immunoproteasome-mediated proteolysis generates immunogenic epitopes presented by major histocompatibility complex (MHC) class I molecules. Threonine at position 75 is highly conserved and its substitution with methionine disrupts the tertiary structure of PSMB8. As compared to normal lymphoblasts, those from an affected patient showed significantly reduced chymotrypsin-like proteolytic activity mediated by immunoproteasomes. We conclude that mutations in PSMB8 cause JMP syndrome, most probably by affecting MHC class I antigen processing.