Recent Advances on Mutant p53: Unveiling Novel Oncogenic Roles, Degradation Pathways, and Therapeutic Interventions.

The p53 protein is the master regulator of cellular integrity, primarily due to its tumor-suppressing functions. Approximately half of all human cancers carry mutations in the TP53 gene, which not only abrogate the tumor-suppressive functions but also confer p53 mutant proteins with oncogenic potential. The latter is achieved through so-called gain-of-function (GOF) mutations that promote cancer progression, metastasis, and therapy resistance by deregulating transcriptional networks, signaling pathways, metabolism, immune surveillance, and cellular compositions of the microenvironment. Despite recent progress in understanding the complexity of mutp53 in neoplastic development, the exact mechanisms of how mutp53 contributes to cancer development and how they escape proteasomal and lysosomal degradation remain only partially understood. In this review, we address recent findings in the field of oncogenic functions of mutp53 specifically regarding, but not limited to, its implications in metabolic pathways, the secretome of cancer cells, the cancer microenvironment, and the regulating scenarios of the aberrant proteasomal degradation. By analyzing proteasomal and lysosomal protein degradation, as well as its connection with autophagy, we propose new therapeutical approaches that aim to destabilize mutp53 proteins and deactivate its oncogenic functions, thereby providing a fundamental basis for further investigation and rational treatment approaches for TP53-mutated cancers.

The deubiquitinase USP2a promotes tumor immunosuppression by stabilizing immune checkpoint B7-H4 in lung adenocarcinoma harboring EGFR-activating mutants.

B7-H4 is an immune checkpoint crucial for inhibiting CD8+ T-cell activity. A clinical trial is underway to investigate B7-H4 as a potential immunotherapeutic agent. However, the regulatory mechanism of B7-H4 degradation via the ubiquitin-proteasome pathway (UPP) remains poorly understood. In this study, we discovered that proteasome inhibitors effectively increased B7-H4 expression, while EGFR-activating mutants promoted B7-H4 expression through the UPP. We screened B7-H4 binding proteins by co-immunoprecipitation and mass spectrometry and found that USP2a acted as a deubiquitinase of B7-H4 by removing K48- and K63-linked ubiquitin chains from B7-H4, leading to a reduction in B7-H4 degradation. EGFR mutants enhanced B7-H4 stability by upregulating USP2a expression. We further investigated the role of USP2a in tumor growth in vivo. Depletion of USP2a in L858R/LLC cells inhibited tumor cell proliferation, consequently suppressing tumor growth in immune-deficient nude mice by destabilizing downstream molecules such as Cyclin D1. In an immune-competent C57BL/6 mouse tumor model, USP2a abrogation facilitated infiltration of CD95+CD8+ effector T cells and hindered infiltration of Tim-3+CD8+ and LAG-3+CD8+ exhausted T cells by destabilizing B7-H4. Clinical lung adenocarcinoma samples showed a significant correlation between B7-H4 abundance and USP2a expression, indicating the contribution of the EGFR/USP2a/B7-H4 axis to tumor immunosuppression. In summary, this study elucidates the dual effects of USP2a in tumor growth by stabilizing Cyclin D1, promoting tumor cell proliferation, and stabilizing B7-H4, contributing to tumor immunosuppression. Therefore, USP2a represents a potential target for tumor therapy.

MG132 dramatically reduces SAA expression in chicken hepatocellular carcinoma cells at the transcript level independent of its endogenous promoter.

BACKGROUND: MG132, a proteasome inhibitor, is widely used to inhibit nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activity by proteasome-mediated degradation of IκB. It has been marketed as a specific, reversible, cell-permeable and low-cost inhibitor. However, adverse effects of the compound have been reported in the literature. We recently discovered and characterised a point mutation in the acute phase protein serum amyloid A (SAA) in chickens, by overexpressing the protein in chicken hepatocellular carcinoma (LMH) cells. This serine to arginine exchange at amino acid position 90 (SAA.R90S) leads to intra- and extracellular accumulation of SAA, which is surprisingly counteracted by MG132 treatment, independent of SAA's intrinsic promoter. METHODS AND RESULTS: To test, whether low proteasomal degradation of SAA.R90S is responsible for the observed intra- and extracellular SAA accumulation, we intended to inhibit the proteasome in SAA wild type (SAA.WT) overexpressing cells with MG132. However, we observed an unexpected drastic decrease in SAA protein expression at the transcript level. NF-κB gene expression was unchanged by MG132 at the measured time point. CONCLUSIONS: The observed results demonstrate that MG132 inhibits SAA expression at the transcript level, independent of its endogenous promoter. Further, the data might indicate that NF-κB is not involved in the observed MG132-induced inhibition of SAA expression. We, consequently, question in this brief report whether MG132 should truly be categorised as a specific ubiquitin proteasome inhibitor and recommend the usage of alternative compounds.

Ubiquitin-mediated regulation of APE2 protein abundance.

APE2 plays important roles in the maintenance of genomic and epigenomic stability including DNA repair and DNA damage response. Accumulating evidence has suggested that APE2 is upregulated in multiple cancers at the protein and mRNA levels and that APE2 upregulation is correlative with higher and lower overall survival of cancer patients depending on tumor type. However, it remains unknown how APE2 protein abundance is maintained and regulated in cells. Here, we provide the first evidence of APE2 regulation via the posttranslational modification ubiquitin. APE2 is poly-ubiquitinated via K48-linked chains and degraded via the ubiquitin-proteasome system where K371 is the key residue within APE2 responsible for its ubiquitination and degradation. We further characterize MKRN3 as the E3 ubiquitin ligase for APE2 ubiquitination in cells and in vitro. In summary, this study offers the first definition of the APE2 proteostasis network and lays the foundation for future studies pertaining to the posttranslational modification regulation and functions of APE2 in genome integrity and cancer etiology/treatment.

Biochemical and functional characterization of the p.A165T missense variant of mitochondrial amidoxime-reducing component 1.

Recent genome-wide association studies have identified a missense variant p.A165T in mitochondrial amidoxime-reducing component 1 (mARC1) that is strongly associated with protection from all-cause cirrhosis and improved prognosis in nonalcoholic steatohepatitis. The precise mechanism of this protective effect is unknown. Substitution of alanine 165 with threonine is predicted to affect mARC1 protein stability and to have deleterious effects on its function. To investigate the mechanism, we have generated a knock-in mutant mARC1 A165T and a catalytically dead mutant C273A (as a control) in human hepatoma HepG2 cells, enabling characterization of protein subcellular distribution, stability, and biochemical functions of the mARC1 mutant protein expressed from its endogenous locus. Compared to WT mARC1, we found that the A165T mutant exhibits significant mislocalization outside of its traditional location anchored in the mitochondrial outer membrane and reduces protein stability, resulting in lower basal levels. We evaluated the involvement of the ubiquitin proteasome system in mARC1 A165T degradation and observed increased ubiquitination and faster degradation of the A165T variant. In addition, we have shown that HepG2 cells carrying the MTARC1 p.A165T variant exhibit lower N-reductive activity on exogenously added amidoxime substrates in vitro. The data from these biochemical and functional assays suggest a mechanism by which the MTARC1 p.A165T variant abrogates enzyme function which may contribute to its protective effect in liver disease.

The Reduction of PSMB4 in T24 and J82 Bladder Cancer Cells Inhibits the Angiogenesis and Migration of Endothelial Cells.

Bladder cancer (BC) is a malignant tumor of the urinary system with high mortality and recurrence rates. Proteasome subunit type 4 (PSMB4) is highly expressed and has been identified as having oncogenic properties in a variety of cancer types. This study aimed to explore the effect of PSMB4 knockdown on the survival, migration, and angiogenesis of human bladder cancer cells with different degrees of malignancy. We analyzed the effects of PSMB4 knockdown in bladder cancer cells and endothelial cells in the tumor microenvironment. PSMB4 was highly expressed in patients with low- and high-grade urothelial carcinoma. Inhibition of PSMB4 reduced protein expression of focal adhesion kinase (FAK) and myosin light chain (MLC), leading to reduced migration. Furthermore, the suppression of PSMB4 decreased the levels of vascular endothelial factor B (VEGF-B), resulting in lower angiogenic abilities in human bladder cancer cells. PSMB4 inhibition affected the migratory ability of HUVECs and reduced VEGFR2 expression, consequently downregulating angiogenesis. In the metastatic animal model, PSMB4 knockdown reduced the relative volumes of lung tumors. Our findings suggest the role of PSMB4 as a potential target for therapeutic strategies against human bladder cancer.

Proteasome gene expression is controlled by coordinated functions of multiple transcription factors.

Proteasome activity is crucial for cellular integrity, but how tissues adjust proteasome content in response to catabolic stimuli is uncertain. Here, we demonstrate that transcriptional coordination by multiple transcription factors is required to increase proteasome content and activate proteolysis in catabolic states. Using denervated mouse muscle as a model system for accelerated proteolysis in vivo, we reveal that a two-phase transcriptional program activates genes encoding proteasome subunits and assembly chaperones to boost an increase in proteasome content. Initially, gene induction is necessary to maintain basal proteasome levels, and in a more delayed phase (7-10 days after denervation), it stimulates proteasome assembly to meet cellular demand for excessive proteolysis. Intriguingly, the transcription factors PAX4 and α-PALNRF-1 control the expression of proteasome among other genes in a combinatorial manner, driving cellular adaptation to muscle denervation. Consequently, PAX4 and α-PALNRF-1 represent new therapeutic targets to inhibit proteolysis in catabolic diseases (e.g., type-2 diabetes, cancer).

TRIM47-CDO1 axis dictates hepatocellular carcinoma progression by modulating ferroptotic cell death through the ubiquitin‒proteasome system.

Hepatocellular carcinoma (HCC) is the predominant form of liver cancer, characterized by high morbidity and mortality rates, as well as unfavorable treatment outcomes. Tripartite motif-containing protein 47 (TRIM47) has been implicated in various diseases including tumor progression with the activity of E3 ubiquitin ligase. However, the precise regulatory mechanisms underlying the involvement of TRIM47 in HCC remain largely unexplored. Here, we provide evidence that TRIM47 exhibits heightened expression in tumor tissues, and its expression is in intimate association with clinical staging and patient prognosis. TRIM47 promotes HCC proliferation, migration, and invasion as an oncogene by in vitro gain- and loss-of-function experiments. TRIM47 knockdown results in HCC ferroptosis induction, primarily through CDO1 involvement to regulate GSH synthesis. Subsequent experiments confirm the interaction between TRIM47 and CDO1 dependent on B30.2 domain, wherein TRIM47 facilitates K48-linked ubiquitination, leading to a decrease in CDO1 protein abundance in HCC. Furthermore, CDO1 is able to counteract the promotional effect of TRIM47 on HCC biological functions. Overall, our research provides novel insight into the mechanism of TRIM47 in CDO1-mediated ferroptosis in HCC cells, highlighting its value as a potential target candidate for HCC therapeutic approaches.

Proteasome Inhibition Reprograms Chromatin Landscape in Breast Cancer.

The 26S proteasome is the major protein degradation machinery in cells. Cancer cells use the proteasome to modulate gene expression networks that promote tumor growth. Proteasome inhibitors have emerged as effective cancer therapeutics, but how they work mechanistically remains unclear. Here, using integrative genomic analysis, we discovered unexpected reprogramming of the chromatin landscape and RNA polymerase II (RNAPII) transcription initiation in breast cancer cells treated with the proteasome inhibitor MG132. The cells acquired dynamic changes in chromatin accessibility at specific genomic loci termed differentially open chromatin regions (DOCR). DOCRs with decreased accessibility were promoter proximal and exhibited unique chromatin architecture associated with divergent RNAPII transcription. Conversely, DOCRs with increased accessibility were primarily distal to transcription start sites and enriched in oncogenic superenhancers predominantly accessible in non-basal breast tumor subtypes. These findings describe the mechanisms by which the proteasome modulates the expression of gene networks intrinsic to breast cancer biology. SIGNIFICANCE: Our study provides a strong basis for understanding the mechanisms by which proteasome inhibitors exert anticancer effects. We find open chromatin regions that change during proteasome inhibition, are typically accessible in non-basal breast cancers.

Disulfide Bonds of Thyroid Peroxidase Are Critical Elements for Subcellular Localization, Proteasome-Dependent Degradation, and Enzyme Activity.

Background: Congenital hypothyroidism (CH) is caused by mutations in cysteine residues, including Cys655 and Cys825 that form disulfide bonds in thyroid peroxidase (TPO). It is highly likely that these disulfide bonds could play an important role in TPO activity. However, to date, no study has comprehensively analyzed cysteine mutations that form disulfide bonds in TPO. In this study, we induced mutations in cysteine residues involved in disulfide bonds formation and analyzed their effect on subcellular localization, degradation, and enzyme activities to evaluate the importance of disulfide bonds in TPO activity. Methods: Vector plasmid TPO mutants, C655F and C825R, known to occur in CH, were transfected into HEK293 cells. TPO activity and protein expression levels were measured by the Amplex red assay and Western blotting. The same procedure was performed in the presence of MG132 proteasome inhibitor. Subcellular localization was determined using immunocytochemistry and flow cytometry. The locations of all disulfide bonds within TPO were predicted using in silico analysis. All TPO mutations associated with disulfide bonds were induced. TPO activity and protein expression levels were also measured in all TPO mutants associated with disulfide bonds using the Amplex red assay and Western blotting. Results: C655F and C825R showed significantly decreased activity and protein expression compared with the wild type (WT) (p < 0.05). In the presence of the MG132 proteasome inhibitor, the protein expression level of TPO increased to a level comparable with that of the WT without increases in its activity. The degree of subcellular distribution of TPO to the cell surface in the mutants was lower compared with the WT TPO. Twenty-four cysteine residues were involved in the formation of 12 disulfide bonds in TPO. All TPO mutants harboring an amino acid substitution in each cysteine showed significantly reduced TPO activity and protein expression levels. Furthermore, the differences in TPO activity depended on the position of the disulfide bond. Conclusions: All 12 disulfide bonds play an important role in the activity of TPO. Furthermore, the mutations lead to misfolding, degradation, and membrane insertion.

The nucleocapsid protein facilitates p53 ubiquitination-dependent proteasomal degradation via recruiting host ubiquitin ligase COP1 in PEDV infection.

Porcine epidemic diarrhea virus (PEDV) is a highly contagious enteric pathogen of the coronavirus family and caused severe economic losses to the global swine industry. Previous studies have established that p53 is a host restriction factor for PEDV infection, and p53 degradation occurs in PEDV-infected cells. However, the underlying molecular mechanisms through which PEDV viral proteins regulate p53 degradation remain unclear. In this study, we found that PEDV infection or expression of the nucleocapsid protein downregulates p53 through a post-translational mechanism: increasing the ubiquitination of p53 and preventing its nuclear translocation. We also show that the PEDV N protein functions by recruiting the E3 ubiquitin ligase COP1 and suppressing COP1 self-ubiquitination and protein degradation, thereby augmenting COP1-mediated degradation of p53. Additionally, COP1 knockdown compromises N-mediated p53 degradation. Functional mapping using truncation analysis showed that the N-terminal domains of N protein were responsible for interacting with COP1 and critical for COP1 stability and p53 degradation. The results presented here suggest the COP1-dependent mechanism for PEDV N protein to abolish p53 activity. This study significantly increases our understanding of PEDV in antagonizing the host antiviral factor p53 and will help initiate novel antiviral strategies against PEDV.

Cisplatin Disrupts Proteasome Bounce-Back Effect through Suppressing ZEB1/Nfe2l1 in Cholangiocarcinoma.

BACKGROUND: Bortezomib (BTZ) is a powerful proteasome inhibitor that has been approved for the treatment of haematologic malignancies. Its effectiveness has been assessed against different types of solid tumours. BTZ is ineffective in most solid tumours because of drug resistance, including cholangiocarcinoma, which is associated with a proteasome bounce-back effect. However, the mechanism through which proteasome inhibitors induce the proteasome bounce-back effect remains largely unknown. METHODS: Cholangiocarcinoma cells were treated with BTZ, cisplatin, or a combination of both. The mRNA levels of Nfe2l1 and proteasome subunit genes (PSMA1, PSMB7, PSMD1, PSMD11, PSMD14, and PSME4) were determined using quantitative real time polymerase chain reaction (qPCR). The protein levels of nuclear factor-erythroid 2-related factor 1 (Nfe2l1) and proteasome enzyme activity were evaluated using western blotting and proteasome activity assays, respectively. Transcriptome sequencing was performed to screen for potential transcription factors that regulate Nfe2l1 expression. The effect of zinc finger E-box-binding homeobox 1 (ZEB1) on the expression of Nfe2l1 and proteasome subunit genes, as well as proteasome enzyme activity, was evaluated after the knockdown of ZEB1 expression with siRNA before treatment with BTZ. The transcriptional activity of ZEB1 on the Nfe2l1 promoter was detected using dual-luciferase reporter gene and chromatin immunoprecipitation assays. Cell viability was measured using the cell counting kit-8 (CCK-8) assay and cell apoptosis was assessed using western blotting and flow cytometry. RESULTS: Cisplatin treatment of BTZ-treated human cholangiocarcinoma cell line (RBE) suppressed proteasome subunit gene expression (proteasome bounce-back) and proteasomal enzyme activity. This effect was achieved by reducing the levels of Nfe2l1 mRNA and protein. Our study utilised transcriptome sequencing to identify ZEB1 as an upstream transcription factor of Nfe2l1, which was confirmed using dual-luciferase reporter gene and chromatin immunoprecipitation assays. Notably, ZEB1 knockdown using siRNA (si-ZEB1) hindered the expression of proteasome subunit genes under both basal and BTZ-induced conditions, leading to the inhibition of proteasomal enzyme activity. Furthermore, the combination treatment with BTZ, cisplatin, and si-ZEB1 significantly reduced the viability of RBE cells. CONCLUSIONS: Our study uncovered a novel mechanism through which cisplatin disrupts the BTZ-induced proteasome bounce-back effect by suppressing the ZEB1/Nfe2l1 axis in cholangiocarcinoma. This finding provides a theoretical basis for developing proteasome inhibitor-based strategies for the clinical treatment of cholangiocarcinoma and other tumours.

PSMB2 plays an oncogenic role in glioma and correlates to the immune microenvironment.

There has been an upward trend in the incidence of glioma, with high recurrence and high mortality. The beta subunits of the 20S proteasome are encoded by the proteasome beta (PSMB) genes and may affect the proteasome's function in glioma, assembly and inhibitor binding. This study attempted to reveal the function of the proliferation and invasion of glioma cells, which is affected by proteasome 20S subunit beta 2 (PSMB2). We subjected the data downloaded from the TCGA database to ROC, survival, and enrichment analyses. After establishing the stable PSMB2 knockdown glioma cell line. We detect the changes in the proliferation, invasion and migration of glioma cells by plate colony formation assay, transwell assay, wound healing assay and flow cytometry and PSMB2 expression was verified by quantitative PCR and Western blotting to identify the mRNA and protein levels. PSMB2 expression was higher in glioma tissues, and its expression positively correlated with poor prognosis and high tumor grade and after PSMB2 knockdown, the proliferation, invasion and migration of glioma cells were weakened.

A tumor suppressor protein encoded by circKEAP1 inhibits osteosarcoma cell stemness and metastasis by promoting vimentin proteasome degradation and activating anti-tumor immunity.

BACKGROUND: Osteosarcoma (OS) is one of most commonly diagnosed bone cancer. Circular RNAs (circRNAs) are a class of highly stable non-coding RNA, the majority of which have not been characterized functionally. The underlying function and molecular mechanisms of circRNAs in OS have not been fully demonstrated. METHOD: Microarray analysis was performed to identify circRNAs that are differentially-expressed between OS and corresponding normal tissues. The biological function of circKEAP1 was confirmed in vitro and in vivo. Mass spectrometry and western blot assays were used to identify the circKEAP1-encoded protein KEAP1-259aa. The molecular mechanism of circKEAP1 was investigated by RNA sequencing and RNA immunoprecipitation analyses. RESULTS: Here, we identified a tumor suppressor circKEAP1, originating from the back-splicing of exon2 of the KEAP1 gene. Clinically, circKEAP1 is downregulated in OS tumors and associated with better survival in cancer patients. N6-methyladenosine (m6A) at a specific adenosine leads to low expression of circKEAP1. Further analysis revealed that circKEAP1 contained a 777 nt long ORF and encoded a truncated protein KEAP1-259aa that reduces cell proliferation, invasion and tumorsphere formation of OS cells. Mechanistically, KEAP1-259aa bound to vimentin in the cytoplasm to promote vimentin proteasome degradation by interacting with the E3 ligase ARIH1. Moreover, circKEAP1 interacted with RIG-I to activate anti-tumor immunity via the IFN-γ pathway. CONCLUSION: Taken together, our findings characterize a tumor suppressor circKEAP1 as a key tumor suppressor regulating of OS cell stemness, proliferation and migration, providing potential therapeutic targets for treatment of OS.

MiR-383 sensitizes osteosarcoma cells to bortezomib treatment via down-regulating PSMB5.

BACKGROUND: Proteasome inhibition is a promising strategy for cancer therapy. Bortezomib, which primarily targets the chymotrypsin-like activity of PSMB5, has demonstrated efficacy in various tumors. However, there is variable sensitivity to bortezomib, which could be attributed, in part, to variations in the expression of proteasome subunits. METHODS AND RESULTS: In this study, we investigated whether miR-383 affects the expression of proteasome subunits in osteosarcoma (OS) cells, and if so, whether OS cells display differential sensitivity to bortezomib concerning miR-383 expression. We detected a decreased miR-383 expression in OS cells and tissues. Then we found a negative correlation between the cytotoxicity of bortezomib and the expression level of the proteasome 20S core particle subunit ß5 (PSMB5). Intriguingly, we identified PSMB5 as a direct target of miR-383. Increased expression of miR-383 resulted in decreased PSMB5 expression and increased sensitivity to bortezomib in OS cells. CONCLUSIONS: In summary, our findings present the initial comprehensive analysis of the function of miR-383 in OS. The outcomes indicate that miR-383 may augment the anticancer effect of bortezomib through PSMB5 repression, offering a novel therapeutic approach in OS and a fresh pathway for proteasome regulation.

Acute accumulation of PIM2 and NRF2 and recovery of ß5 subunit activity mitigate multiple myeloma cell susceptibility to proteasome inhibitors.

Resistance to proteasome inhibitors (PIs) has emerged as an important clinical issue. We investigated the mechanisms underlying multiple myeloma (MM) cell resistance to PIs. To mimic their pharmacokinetic/pharmacodynamic (PK/PD) profiles, MM cells were treated with bortezomib and carfilzomib for 1 h at concentrations up to 400 and 1,000 nM, respectively. Susceptibility to these PIs markedly varied among MM cell lines. Pulsatile treatments with PIs suppressed translation, as demonstrated by incorporation of puromycin at 24 h in PI-susceptible MM.1S cells, but not PI-resistant KMS-11 cells. Inhibition of ß5 subunit activity decreased at 24 h in KMS-11 cells, even with the irreversible PI carfilzomib, but not under suppression of protein synthesis with cycloheximide. Furthermore, the proteasome-degradable pro-survival factors PIM2 and NRF2 acutely accumulated in MM cells subjected to pulsatile PI treatments. Accumulated NRF2 was trans-localized into the nucleus to induce the expression of its target gene, HMOX1, in MM cells. PIM and Akt inhibition restored the anti-MM effects of PIs, even against PI-resistant KMS-11 cells. Collectively, these results suggest that increased synthesis of ß5 proteasome subunit and acute accumulation of PIM2 and NRF2 reduce the anti-MM effects of PIs.

Transcription factor PbNAC71 regulates xylem and vessel development to control plant height.

Dwarfism is an important agronomic trait in fruit breeding programs. However, the germplasm resources required to generate dwarf pear (Pyrus spp.) varieties are limited. Moreover, the mechanisms underlying dwarfism remain unclear. In this study, "Yunnan" quince (Cydonia oblonga Mill.) had a dwarfing effect on "Zaosu" pear. Additionally, the dwarfism-related NAC transcription factor gene PbNAC71 was isolated from pear trees comprising "Zaosu" (scion) grafted onto "Yunnan" quince (rootstock). Transgenic Nicotiana benthamiana and pear OHF-333 (Pyrus communis) plants overexpressing PbNAC71 exhibited dwarfism, with a substantially smaller xylem and vessel area relative to the wild-type controls. Yeast one-hybrid, dual-luciferase, chromatin immunoprecipitation-qPCR, and electrophoretic mobility shift assays indicated that PbNAC71 downregulates PbWalls are thin 1 expression by binding to NAC-binding elements in its promoter. Yeast two-hybrid assays showed that PbNAC71 interacts with the E3 ubiquitin ligase PbRING finger protein 217 (PbRNF217). Furthermore, PbRNF217 promotes the ubiquitin-mediated degradation of PbNAC71 by the 26S proteasome, thereby regulating plant height as well as xylem and vessel development. Our findings reveal a mechanism underlying pear dwarfism and expand our understanding of the molecular basis of dwarfism in woody plants.

Development of an ubiquitin-proteasome system signature for predicting prognosis and providing therapeutic guidance for patients with triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is a pathological subtype with a high mortality, and the development of inhibitors in the ubiquitin-proteasome system (UPS) component could be a novel therapeutic tool. METHODS: Triple-negative breast cancer data were obtained from The Cancer Genome Atlas (TCGA), and subtype analysis was performed by consistent clustering analysis to identify molecular subtypes of TNBC according to UPS characteristics. Differential analysis, COX and least absolute shrinkage and selection operator (LASSO) COX regression analyses were performed to select genes associated with overall survival in TNBC. The final prognostic model (UPS score) was determined using the LASSO COX model. The model performance was assessed using receiver operating characteristic (ROC) curves and survival curves. In addition, the results of the UPS score on analyzing the abundance of immune cell infiltration and immunotherapy were explored. Finally, we developed a nomogram for TNBC survival prediction. RESULTS: Two UPS subtypes (UPSMS1 and UPSMS2) showing significant survival differences were classified. COX regression analysis on differentially expressed genes in UPSMS1 and UPSMS2 filtered five genes that affected overall survival. Based on the regression coefficients and expression data of the five genes, we built a prognostic assessment system (UPS score). The UPS score showed consistent prognostic and therapeutic guidance values. Finally, the ROC curve of the nomogram and UPS score showed the highest predictive efficacy compared with traditional clinical prognostic indicators. CONCLUSION: The UPS score represented a promising prognostic tool to predict overall survival and immune status and guide personalized treatment selection in TNBC patients, and this study may provide a more practical alternative for clinical monitoring and management of TNBC.

LncRNA PSMB8-AS1 Instigates Vascular Inflammation to Aggravate Atherosclerosis.

BACKGROUND: Increasing evidence suggests that long noncoding RNAs play significant roles in vascular biology and disease development. One such long noncoding RNA, PSMB8-AS1, has been implicated in the development of tumors. Nevertheless, the precise role of PSMB8-AS1 in cardiovascular diseases, particularly atherosclerosis, has not been thoroughly elucidated. Thus, the primary aim of this investigation is to assess the influence of PSMB8-AS1 on vascular inflammation and the initiation of atherosclerosis. METHODS: We generated PSMB8-AS1 knockin and Apoe (Apolipoprotein E) knockout mice (Apoe-/-PSMB8-AS1KI) and global Apoe and proteasome subunit-ß type-9 (Psmb9) double knockout mice (Apoe-/-Psmb9-/-). To explore the roles of PSMB8-AS1 and Psmb9 in atherosclerosis, we fed the mice with a Western diet for 12 weeks. RESULTS: Long noncoding RNA PSMB8-AS1 is significantly elevated in human atherosclerotic plaques. Strikingly, Apoe-/-PSMB8-AS1KI mice exhibited increased atherosclerosis development, plaque vulnerability, and vascular inflammation compared with Apoe-/- mice. Moreover, the levels of VCAM1 (vascular adhesion molecule 1) and ICAM1 (intracellular adhesion molecule 1) were significantly upregulated in atherosclerotic lesions and serum of Apoe-/-PSMB8-AS1KI mice. Consistently, in vitro gain- and loss-of-function studies demonstrated that PSMB8-AS1 induced monocyte/macrophage adhesion to endothelial cells and increased VCAM1 and ICAM1 levels in a PSMB9-dependent manner. Mechanistic studies revealed that PSMB8-AS1 induced PSMB9 transcription by recruiting the transcription factor NONO (non-POU domain-containing octamer-binding protein) and binding to the PSMB9 promoter. PSMB9 (proteasome subunit-ß type-9) elevated VCAM1 and ICAM1 expression via the upregulation of ZEB1 (zinc finger E-box-binding homeobox 1). Psmb9 deficiency decreased atherosclerotic lesion size, plaque vulnerability, and vascular inflammation in Apoe-/- mice in vivo. Importantly, endothelial overexpression of PSMB8-AS1-increased atherosclerosis and vascular inflammation were attenuated by Psmb9 knockout. CONCLUSIONS: PSMB8-AS1 promotes vascular inflammation and atherosclerosis via the NONO/PSMB9/ZEB1 axis. Our findings support the development of new long noncoding RNA-based strategies to counteract atherosclerotic cardiovascular disease.

PSMD14 stabilizes estrogen signaling and facilitates breast cancer progression via deubiquitinating ERα.

The over-activation of ERα signaling is regarded as the major driver for luminal breast cancers, which could be effective controlled via selective estrogen receptor modulators (SERM), such as tamoxifen. The endocrine resistance is still a challenge for breast cancer treatment, while recently studies implicate the post-translational modification on ERα play important roles in endocrine resistance. The stability of ERα protein and ERα transcriptome are subject to a balance between E3 ubiquitin ligases and deubiquitinases. Through deubiquitinases siRNA library screening, we discover PSMD14 as a critical deubiquitinase for ERα signaling and breast cancer progression. PSMD14 could facilitate breast cancer progression through ERα signaling in vitro and in vivo, while pharmaceutical inhibition of PSMD14 via Thiolutin could block the tumorigenesis in breast cancer. In endocrine resistant models, PSMD14 inhibition could de-stabilize the resistant form of ERα (Y537S) and restore tamoxifen sensitivity. Molecular studies reveal that PSMD14 could inhibition K48-linked poly-ubiquitination on ERα, facilitate ERα transcriptome. Interestingly, ChIP assay shows that ERα could bind to the promoter region of PSMD14 and facilitate its gene transcription, which indicates PSMD14 is both the upstream modulator and downstream target for ERα signaling in breast cancer. In general, we identified a novel positive feedback loop between PSMD14 and ERα signaling in breast cancer progression, while blockade of PSMD14 could be a plausible strategy for luminal breast cancer.