RESUMO
Glioma is primary brain tumour with a poor prognosis. Metabolic reprogramming is a hallmark of glioma, and is critical in the development of antiglioma agents and glioma therapy. Cuproptosis is a novel form of cell death mediated by protein lipidation and highly associated with mitochondrial metabolism. However, the clinical impact of cuproptosis-related genes (CRGs) in glioma remains largely unknown. The purpose of this study is to create a new CRGs signature that can be used to predict survival and immunotherapy in glioma patients. LASSO regression analysis was applied to establish prognostic gene signatures. Furthermore, a CRGs signature-based nomogram was developed and demonstrated good predictive potential. We also analyzed the relationship of CRGs and immune infiltration and the correlation with the pathological grade of glioma. Finally, we explored the miRNA that may regulate cuproptosis-related gene FDX1. We found that miR-606 was markedly downregulated in GBM, overexpression of miR-606 can significantly inhibit aerobic glycolysis and proliferation of GBM cells. FDX1 was upregulated in GBM, knockdown of FDX1 significantly inhibit aerobic glycolysis and proliferation of GBM cells. And luciferase assay was used to verified that miR-606 binds to and regulates FDX1 mRNA. These results provide a basis for further exploring the biological mechanisms of cuproptosis. This study may provide new potential therapeutic perspectives for patients with glioma.
Assuntos
Apoptose , Glioma , MicroRNAs , Humanos , Morte Celular , Cobre , Glioma/genética , MicroRNAs/genética , Nomogramas , PrognósticoRESUMO
Glioblastoma (GBM) is a primary tumor in the intracranial compartment. Vasculogenic mimicry (VM) is a process in which a pipeline of tumor cells that provide blood support to carcinogenic cells is formed, and studying VM could provide a new strategy for clinical targeted treatment of GBM. In the present study, we found that SNORD17 and ZNF384 were significantly upregulated and promoted VM in GBM, whereas KAT6B was downregulated and inhibited VM in GBM. RTL-P assays were performed to verify the 2'-O-methylation of KAT6B by SNORD17; IP assays were used to detect the acetylation of ZNF384 by KAT6B. In addition, the binding of ZNF384 to the promoter regions of VEGFR2 and VE-cadherin promoted transcription, as validated by chromatin immunoprecipitation and luciferase reporter assays. And finally, knockdown of SNORD17 and ZNF384 combined with KAT6B overexpression effectively reduced the xenograft tumor size, prolonged the survival time of nude mice and reduced the number of VM channels. This study reveals a novel mechanism of the SNORD17/KAT6B/ZNF384 axis in modulating VM development in GBM that may provide a new goal for the comprehensive treatment of GBM.
Assuntos
Glioblastoma , Animais , Camundongos , Humanos , Glioblastoma/genética , Glioblastoma/tratamento farmacológico , Camundongos Nus , Metilação , Linhagem Celular Tumoral , RNA Mensageiro , Histona Acetiltransferases/uso terapêuticoRESUMO
RNA-binding proteins (RBPs), long non-coding RNAs (lncRNAs), and small nucleolar RNAs (snoRNAs) were found to play crucial regulatory roles in ischemic injury. Based on GEO databases and our experimental results, we selected Dcp2, lncRNA-RNCR3, Dkc1, and Snora62 and Foxh1 as research candidates. We found that expression levels of Dcp2, RNCR3, Dkc1, Snora62, and Foxh1 were upregulated in oxygen glucose deprivation-treated HT22 cells and hippocampal tissues subject to chronic cerebral ischemia (CCI). Silencing of Dcp2, RNCR3, Dkc1, Snora62, and Foxh1 all inhibited apoptosis of oxygen glucose deprivation-treated HT22 cells. Moreover, Dcp2 promoted RNCR3 expression by increasing its stability. Importantly, RNCR3 may act as a molecular skeleton to bind to Dkc1 and recruit Dck1 to promote snoRNP assembly. Snora62 was responsible for pseudouridylation at 28S rRNA U3507 and U3509 sites. Pseudouridylation levels of 28S rRNA were reduced after knockdown of Snora62. Decreased pseudouridylation levels inhibited the translational activity of its downstream target, Foxh1. Our study further confirmed that Foxh1 transcriptionally promoted the expression of Bax and Fam162a. Notably, experiments in vivo showed that Dcp2 knockdown combined with RNCR3 knockdown and Snora62 knockdown resulted in an anti-apoptosis effect. In conclusion, this study suggests that the axis Dcp2/RNCR3/Dkc1/Snora621 is important for the regulation of neuronal apoptosis induced by CCI.
Assuntos
Isquemia Encefálica , MicroRNAs , RNA Longo não Codificante , Humanos , MicroRNAs/genética , RNA Ribossômico 28S , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Apoptose/genética , Glucose , Oxigênio , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismoRESUMO
Human malignant gliomas are the most common and aggressive primary malignant tumors of the human central nervous system. Vasculogenic mimicry (VM), which refers to the formation of a tumor blood supply system independently of endothelial cells, contributes to the malignant progression of glioma. Therefore, VM is considered a potential target for glioma therapy. Accumulated evidence indicates that alterations in SUMOylation, a reversible post-translational modification, are involved in tumorigenesis and progression. In the present study, we found that UBA2 and RALY were upregulated in glioma tissues and cell lines. Downregulation of UBA2 and RALY inhibited the migration, invasion, and VM of glioma cells. RALY can be SUMOylated by conjugation with SUMO1, which is facilitated by the overexpression of UBA2. The SUMOylation of RALY increases its stability, which in turn increases its expression as well as its promoting effect on FOXD1 mRNA. The overexpression of FOXD1 promotes DKK1 transcription by activating its promoter, thereby promoting glioma cell migration, invasion, and VM. Remarkably, the combined knockdown of UBA2, RALY, and FOXD1 resulted in the smallest tumor volumes and the longest survivals of nude mice in vivo. UBA2/RALY/FOXD1/DKK1 axis may play crucial roles in regulating VM in glioma, which may contribute to the development of potential strategies for the treatment of gliomas.
Assuntos
Neoplasias Encefálicas , Glioma , Camundongos , Animais , Humanos , Neoplasias Encefálicas/metabolismo , Sumoilação , Camundongos Nus , Células Endoteliais/metabolismo , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Glioma/genética , Glioma/metabolismo , Linhagem Celular Tumoral , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/metabolismo , Enzimas Ativadoras de Ubiquitina/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Fatores de Transcrição Forkhead/genéticaRESUMO
RNA-binding proteins (RBPs) are significantly dysregulated in glioma. In this study, we demonstrated the upregulation of Nuclear cap-binding subunit 3 (NCBP3) in glioma tissues and cells. Further, knockdown of NCBP3 inhibited the malignant progression of glioma. NCBP3 directly bound to small nucleolar RNA host gene 6 (SNHG6) and stabilized SNHG6 expression. In contrast, the gastrulation brain homeobox 2 (GBX2) transcription factor was downregulated in glioma tissues and cells. SNHG6 inhibited GBX2 transcription by mediating the H3K27me3 modification induced by polycomb repressive complex 2 (PRC2). Moreover, GBX2 decreased the promoter activities and downregulated the expression of the flotillin protein family 1 (FLOT1) oncogene. In conclusion, NCBP3/SNHG6 inhibits GBX2 transcription in a PRC2-dependent manner to facilitate the malignant progression of gliomas.
Assuntos
Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/metabolismo , Histonas/metabolismo , Proteínas de Homeodomínio/genética , Interferência de RNA , RNA Longo não Codificante/genética , Linhagem Celular Tumoral , Progressão da Doença , Técnicas de Silenciamento de Genes , Glioma/patologia , Humanos , Gradação de Tumores , Estadiamento de Neoplasias , Regiões Promotoras Genéticas , Ligação Proteica , Transcrição Gênica , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
Studies have found that RNA-binding proteins (RBPs) and long non-coding RNAs (lncRNAs) are dysregulated and play an important regulatory role in the development of tumors. Based on The Cancer Genome Atlas (TCGA) database, our findings from experiments, and the evidence of previous studies, we screened DiGeorge syndrome critical region gene 8 (DGCR8), ZFAT antisense RNA 1 (ZFAT-AS1), and caudal type homeobox 2 (CDX2) as research candidates. In the present study, DGCR8 and CDX2 were highly expressed and ZFAT-AS1 was markedly downregulated in glioma tissues and cells. DGCR8 or CDX2 knockdown or ZFAT-AS1 overexpression suppressed glioma cell proliferation, migration, and invasion and facilitated apoptosis. DGCR8 might decrease ZFAT-AS1 expression by attenuating its stability in a manner of inducing its cleavage. Importantly, ZFAT-AS1 could inhibit CDX2 transcription by mediating the methylation of histone H3 on lysine 27 (H3K27me3) modification induced by PRC2 in the CDX2 promoter region. In addition, CDX2 transcriptionally activated DGCR8 expression by binding to its promoter regions, forming a positive feedback loop of DGCR8/ZFAT-AS1/CDX2. In conclusion, DGCR8/ZFAT-AS1 promotes CDX2 transcription in a PRC2 complex-dependent manner to facilitate the malignant biological behavior of glioma cells.
Assuntos
Fator de Transcrição CDX2/genética , Regulação Neoplásica da Expressão Gênica , Glioma/genética , RNA Antissenso , RNA Longo não Codificante/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Modelos Animais de Doenças , Técnicas de Silenciamento de Genes , Glioma/metabolismo , Glioma/mortalidade , Glioma/patologia , Humanos , Camundongos , Regiões Promotoras Genéticas , Ligação Proteica , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Increasing evidence has suggested that gliomas can supply blood through vasculogenic mimicry. In this study, the expression and function of ZNRD1-AS1-144aa-uORF (144aa-uORF) and some non-coding RNAs in gliomas were assessed. Real-time quantitative PCR or Western blot was used to discover the expression of 144aa-uORF, ZNRD1-AS1, miR-499a-5p, ELF1 and EMI1 in gliomas. In addition, RIP and RNA pull-down assays were applied to explore the interrelationship between 144aa-uORF and ZNRD1-AS1. The role of the 144aa-uORF\ZNRD1-AS1\miR-499a-5p\ELF1\EMI1 axis in vasculogenic mimicry formation of gliomas was analysed. This study illustrates the reduced expression of the 144aa-uORF in glioma tissues and cells. Up-regulation of 144aa-uORF inhibits proliferation, migration, invasion and vasculogenic mimicry formation within glioma cells. The up-regulated 144aa-uORF can increase the degradation of ZNRD1-AS1 through the nonsense-mediated RNA decay (NMD) pathway. Knockdown of ZNRD1-AS1 inhibits vasculogenic mimicry in glioma cells by modulating miR-499a-5p. At the same time, miR-499a-5p is down-regulated and has a tumour-suppressive effect in gliomas. In addition, ZNRD1-AS1 serves as a competitive endogenous RNA (ceRNA) and regulates the expression of ELF1 by binding to miR-499a-5p. Notably, ELF1 binds to the promoter region of EMI1 and up-regulates EMI1 expression, while simultaneously promoting vasculogenic mimicry in glioma cells. This study suggests that the 144aa-uORF\ZNRD1-AS1\miR-499a-5p\ELF1\EMI1 axis takes key part in regulating the formation of vasculogenic mimicry in gliomas and may provide a potential target for glioma treatment.
Assuntos
Neoplasias Encefálicas/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas F-Box/metabolismo , Glioma/genética , Antígenos de Histocompatibilidade Classe I/metabolismo , MicroRNAs/metabolismo , Proteínas Nucleares/metabolismo , Fases de Leitura Aberta/genética , Transdução de Sinais , Fatores de Transcrição/metabolismo , Animais , Sequência de Bases , Ligação Competitiva , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo/genética , Regulação Neoplásica da Expressão Gênica , Glioma/patologia , Células HEK293 , Humanos , Camundongos Nus , MicroRNAs/genética , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética , Estabilidade de RNA/genética , Análise de Sobrevida , Regulação para Cima/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The blood-brain barrier (BBB) plays a pivotal role in the maintenance and regulation of the neural microenvironment. The BBB breakdown is a pathological change in early Alzheimer's disease (AD). RNA-binding proteins (RBPs) and long non-coding RNAs (lncRNAs) are involved in the regulation of BBB permeability. Our study demonstrates the role of TRA2A/LINC00662/ELK4 axis in regulating BBB permeability in AD microenvironment. In Aß1-42-incubated microvascular endothelial cells (ECs) of the BBB model in vitro, TRA2A and LINC00662 were enriched. TRA2A increased the stability of LINC00662 by binding with it. The knockdown of either TRA2A or LINC00662 decreased BBB permeability due to increased expression of tight junction-related proteins. ELK4 was less expressed in the BBB model in AD microenvironment in vitro. LINC00662 mediated the degradation of ELK4 mRNA by SMD pathway. Downregulation of ELK4 increased BBB permeability by increasing the tight junction-related protein expression.TRA2A/LINC00662/ELK4 axis plays a crucial role in the regulation of BBB permeability in AD microenvironment, which may provide a novel target for the therapy of AD.
Assuntos
Barreira Hematoencefálica/metabolismo , Microambiente Celular/genética , Regulação da Expressão Gênica , RNA Longo não Codificante/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Elk-4 do Domínio ets/genética , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Biomarcadores , Células Endoteliais/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Fragmentos de Peptídeos/metabolismo , Permeabilidade , Regiões Promotoras Genéticas , Ligação Proteica , Interferência de RNA , Estabilidade de RNA , RNA Longo não Codificante/genética , Proteínas de Junções Íntimas/genética , Proteínas de Junções Íntimas/metabolismoRESUMO
Blood-tumour barrier (BTB) has been known to significantly attenuate the efficacy of chemotherapy for glioma. In this report, we identified that insulin-like grown factor 2 mRNA-binding protein 2 (IGF2BP2) was over-expressed in glioma microvessel and glioma endothelial cells (GECs). Knockdown of IGF2BP2 decreased the expression of lncRNA FBXL19-AS1 and tight junction-related proteins, thereby promoting BTB permeability. FBXL19-AS1 was over-expressed and more enriched in the cytoplasm of GECs. In addition, FBXL19-AS1 could bind to 3'-UTR of ZNF765 mRNA and down-regulate ZNF765 mRNA expression through STAU1-mediated mRNA decay (SMD). The low expression of ZNF765 was discovered in GECs and verified to increase BTB permeability by inhibiting the promoter activities of tight junction-related proteins. Meanwhile, ZNF765 also inhibited the transcriptional activity of IGF2BP2, thereby forming a feedback loop in regulating the BTB permeability. Single or combined application of silenced IGF2BP2 and FBXL19-AS1 improved the delivery and antitumor efficiency of doxorubicin (DOX). In general, our study revealed the regulation mechanism of IGF2BP2/FBXL19-AS1/ZNF765 axis on BTB permeability, which may provide valuable insight into treatment strategy for glioma.
Assuntos
Proteínas do Citoesqueleto/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas F-Box/genética , Regulação Neoplásica da Expressão Gênica , RNA Longo não Codificante/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Microambiente Tumoral , Linhagem Celular Tumoral , Células Endoteliais/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Degradação do RNAm Mediada por Códon sem Sentido , Permeabilidade , Ligação Proteica , Estabilidade de RNA , Transcriptoma , Microambiente Tumoral/genéticaRESUMO
Dysregulation of long non-coding RNAs (lncRNAs) confirm that it plays a crucial role in tumourigenesis and malignant progression of glioma. The present study demonstrated that LncRNA secretory carrier membrane protein 1 (SCAMP1) was up-regulated and functioned as an oncogene in glioma cells. In addition, miR-499a-5p was down-regulated meanwhile exerted tumour-suppressive function in glioma cells. Subsequently, inhibition of SCAMP1 significantly restrained the cell proliferation, migration and invasion, as well as promoted apoptosis by acting as a molecular sponge of miR-499a-5p. Transcription factor LIM homeobox transcription factor 1, alpha (LMX1A) was overexpressed in glioma tissues and cells. Moreover, miR-499a-5p targeted LMX1A 3'-UTR in a sequence-specific manner. Hence, down-regulation of SCAMP1 remarkably reduced the expression level of LMX1A, indicating that LMX1A participated in miR-499a-5p-induced tumour-suppressive effects on glioma cells. Furthermore, knockdown of LMX1A decreased NLR family, CARD domain containing 5 (NLRC5) mRNA and protein expression levels through directly binding to the NLRC5 promoter region. Down-regulation of NLRC5 obviously inhibited malignant biological behaviours of glioma cells through attenuating the activity of Wnt/ß-catenin signalling pathway. In conclusion, our study clarifies that SCAMP1/miR-499a-5p/LMX1A/NLRC5 axis plays a critical role in modulating malignant progression of glioma cells, which provide a novel therapeutic strategy for glioma treatment.
Assuntos
Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Homeodomínio LIM/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Fatores de Transcrição/genética , Proteínas de Transporte Vesicular/metabolismo , Regiões 3' não Traduzidas , Animais , Apoptose/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Imunoprecipitação da Cromatina , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Homeodomínio LIM/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Análise de Sequência com Séries de Oligonucleotídeos , Prognóstico , Regiões Promotoras Genéticas , RNA Longo não Codificante/genética , Fatores de Transcrição/metabolismo , Proteínas de Transporte Vesicular/genética , Via de Sinalização Wnt/genéticaRESUMO
Glioma is the most common primary brain tumor, and targeting glioma stem cells (GSCs) has become a key aspect of glioma treatment. In this study, we discovered a molecular network in which circRNA forms an R-loop structure with its parental gene to regulate the biological behavior of GSCs. Genes with abnormal expression in GSCs were screened using RNA-seq and circRNA microarray analyses. The study results showed that high expression of YTHDC1 in GSCs promoted the transportation of N6-methyladenosine (m6A)-modified circPOLR2B from the nucleus to the cytoplasm. Decreased circPOLR2B levels in the nucleus resulted in fewer R-loop structures formed with its parental gene POLR2B. This reduction in R-loop structures relieved the inhibitory effect on POLR2B transcription and upregulated PBX1 expression through alternative polyadenylation (APA) action, thereby promoting the malignant biological behavior of GSCs. Knockdown of YTHDC1, POLR2B, and PBX1 reduced xenograft tumor volume and prolonged the survival of nude mice. The YTHDC1/circPOLR2B/POLR2B/PBX1 axis plays a regulatory role in the biological behavior of GSCs, offering potential targets and novel strategies for the treatment of glioma.
Assuntos
Glioma , Células-Tronco Neoplásicas , RNA Circular , Animais , Humanos , Camundongos , Adenosina/análogos & derivados , Adenosina/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Retroalimentação Fisiológica , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Glioma/patologia , Glioma/metabolismo , Metilação , Camundongos Endogâmicos BALB C , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas do Tecido Nervoso , Fatores de Processamento de RNA/metabolismo , Fatores de Processamento de RNA/genética , RNA Circular/genética , RNA Circular/metabolismoRESUMO
Chronic cerebral ischemia (CCI) is a clinical syndrome characterised by brain dysfunction due to decreased chronic cerebral perfusion. CCI initiates several inflammatory pathways, including pyroptosis. RNA-binding proteins (RBPs) play important roles in CCI. This study aimed to explore whether the interaction between RBP-Cpeb4 and Dclk2 affected Ehf phosphorylation to regulate neuronal pyroptosis. HT22 cells and mice were used to construct oxygen glucose deprivation (OGD)/CCI models. We found that Cpeb4 and Dclk2 were upregulated in OGD-treated HT22 cells and CCI-induced hippocampal CA1 tissues. Cpeb4 upregulated Dclk2 expression by increasing Dclk2 mRNA stability. Knockdown of Cpeb4 or Dclk2 inhibited neuronal pyroptosis in OGD-treated HT22 cells and CCI-induced hippocampal CA1 tissues. By binding to the promoter regions of Caspase1 and Caspase3, the transcription factor Ehf reduced their promoter activities and inhibited the transcription. Dclk2 phosphorylated Ehf and changed its nucleoplasmic distribution, resulting in the exit of p-Ehf from the nucleus and decreased Ehf levels. It promoted the expression of Caspase1 and Caspase3 and stimulated neuronal pyroptosis of HT22 cells induced by OGD. Cpeb4/Dclk2/Ehf pathway plays an important role in the regulation of cerebral ischemia-induced neuronal pyroptosis.
RESUMO
Angiogenesis plays a major role in tumor initiation, progression, and metastasis. This is why finding antiangiogenic targets is essential in the treatment of gliomas. In this study, NSUN2 and LINC00324 were significantly upregulated in conditionally cultured glioblastoma endothelial cells (GECs). Knockdown of NSUN2 or LINC00324 inhibits GECs angiogenesis. NSUN2 increased the stability of LINC00324 by m5C modification and upregulated LINC00324 expression. LINC00324 competes with the 3'UTR of CBX3 mRNA to bind to AUH protein, reducing the degradation of CBX3 mRNA. In addition, CBX3 directly binds to the promoter region of VEGFR2, enhances VEGFR2 transcription, and promotes GECs angiogenesis. These findings demonstrated NSUN2/LINC00324/CBX3 axis plays a crucial role in regulating glioma angiogenesis, which provides new strategies for glioma therapy.
Assuntos
Células Endoteliais , Glioma , Humanos , Células Endoteliais/metabolismo , Angiogênese , Proliferação de Células/genética , Glioma/patologia , RNA Mensageiro/genética , Proteínas Cromossômicas não HistonaRESUMO
Glioblastoma (GBM) is the most common malignant tumor of the central nervous system. Vasculogenic mimicry (VM) is a hematological system composed of tumor cells that exert blood perfusion without relying on vascular endothelial cells. The current poor results of anti-vascular therapy for clinical GBM are associated with the presence of VM; therefore, it is important to investigate VM formation in GBM. Our results demonstrate that STK24P1 encodes P1-121aa with a kinase structural domain, and in vitro kinase assays demonstrated that P1-121aa mediates modification of ELF2 phosphorylation. ChIP and dual luciferase reporter gene assays demonstrated that the transcription factor ELF2 binds to VE-cadherin and the VEGFR2 promoter region, thereby promoting VM formation in glioma cells. P1-121aa, encoded by the pseudogene STK24P1, phosphorylates ELF2 at S107, increasing the stability of the ELF2 protein. ELF2 promotes VEGFR2 and VE-cadherin expression at the transcriptional level, which in turn promotes VM in GBM. This study demonstrates the important roles of STK24P1, P1-121aa, and ELF2 in regulating VM in GBM, which could provide potential targets for GBM treatment.
Assuntos
Glioblastoma , Humanos , Glioblastoma/tratamento farmacológico , Fosforilação , Células Endoteliais/metabolismo , Neovascularização Patológica/tratamento farmacológico , Peptídeos/metabolismo , Linhagem Celular Tumoral , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Glioma is the most common primary malignancy of the central nervous system. Glioblastoma (GBM) has the highest degree of malignancy among the gliomas and the strongest resistance to chemotherapy and radiotherapy. Vasculogenic mimicry (VM) provides tumor cells with a blood supply independent of endothelial cells and greatly restricts the therapeutic effect of anti-angiogenic tumor therapy for glioma patients. Vascular endothelial growth factor receptor 2 (VEGFR2) and vascular endothelial cadherin (VE-cadherin) are currently recognized molecular markers of VM in tumors. In the present study, we show that pseudogene MAPK6P4 deficiency represses VEGFR2 and VE-cadherin protein expression levels, as well as inhibits the proliferation, migration, invasion, and VM development of GBM cells. The MAPK6P4-encoded functional peptide P4-135aa phosphorylates KLF15 at the S238 site, promoting KLF15 protein stability and nuclear entry to promote GBM VM formation. KLF15 was further confirmed as a transcriptional activator of LDHA, where LDHA binds and promotes VEGFR2 and VE-cadherin lactylation, thereby increasing their protein expression. Finally, we used orthotopic and subcutaneous xenografted nude mouse models of GBM to verify the inhibitory effect of the above factors on GBM VM development. In summary, this study may represent new targets for the comprehensive treatment of glioma.
Assuntos
Glioblastoma , Glioma , Animais , Camundongos , Humanos , Glioblastoma/genética , Glioblastoma/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Células Endoteliais/metabolismo , Pseudogenes , Linhagem Celular Tumoral , Glioma/patologiaRESUMO
AIMS: The isocitrate dehydrogenase (IDH) phenotype is associated with reprogrammed energy metabolism in glioblastoma (GBM) cells. Small nucleolar RNAs (snoRNAs) are known to exert an important regulatory role in the energy metabolism of tumor cells. The purpose of this study was to investigate the role of C/D box snoRNA U3 and transcription factor zinc finger and BTB domain-containing 7A (ZBTB7A) in the regulation of aerobic glycolysis and the proliferative capacity of IDH1 wild-type (IDH1WT ) GBM cells. METHODS: Quantitative reverse transcription PCR and western blot assays were utilized to detect snoRNA U3 and ZBTB7A expression. U3 promoter methylation status was analyzed via bisulfite sequencing and methylation-specific PCR. Seahorse XF glycolysis stress assays, lactate production and glucose consumption measurement assays, and cell viability assays were utilized to detect glycolysis and proliferation of IDH1WT GBM cells. RESULTS: We found that hypomethylation of the CpG island in the promoter region of U3 led to the upregulation of U3 expression in IDH1WT GBM cells, and the knockdown of U3 suppressed aerobic glycolysis and the proliferation ability of IDH1WT GBM cells. We found that small nucleolar-derived RNA (sdRNA) U3-miR, a small fragment produced by U3, was able to bind to the ZBTB4 3'UTR region and reduce ZBTB7A mRNA stability, thereby downregulating ZBTB7A protein expression. Furthermore, ZBTB7A transcriptionally inhibited the expression of hexokinase 2 (HK2) and lactate dehydrogenase A (LDHA), which are key enzymes of aerobic glycolysis, by directly binding to the HK2 and LDHA promoter regions, thereby forming the U3/ZBTB7A/HK2 LDHA pathway that regulates aerobic glycolysis and proliferation of IDH1WT GBM cells. CONCLUSION: U3 enhances aerobic glycolysis and proliferation in IDH1WT GBM cells via the U3/ZBTB7A/HK2 LDHA axis.
Assuntos
Glioblastoma , Humanos , Glioblastoma/patologia , RNA Nucleolar Pequeno/metabolismo , Isocitrato Desidrogenase/genética , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Fatores de Transcrição/genética , Glicólise , Proliferação de Células/genética , Regulação Neoplásica da Expressão GênicaRESUMO
Glioblastoma multiforme (GBM) is a highly vascularized malignant cancer of the central nervous system, and the presence of vasculogenic mimicry (VM) severely limits the effectiveness of anti-vascular therapy. In this study, we identified downregulated circHECTD1, which acted as a key VM-suppressed factor in GBM. circHECTD1 elevation significantly inhibited cell proliferation, migration, invasion and tube-like structure formation in GBM. RIP assay was used to demonstrate that the flanking intron sequence of circHECTD1 can be specifically bound by RBMS3, thereby inducing circHECTD1 formation to regulate VM formation in GBM. circHECTD1 was confirmed to possess a strong protein-encoding capacity and the encoded functional peptide 463aa was identified by LC-MS/MS. Both circHECTD1 and 463aa significantly inhibited GBM VM formation in vivo and in vitro. Analysis of the 463aa protein sequence revealed that it contained a ubiquitination-related domain and promoted NR2F1 degradation by regulating the ubiquitination of the NR2F1 at K396. ChIP assay verified that NR2F1 could directly bind to the promoter region of MMP2, MMP9 and VE-cadherin, transcriptionally promoting the expression of VM-related proteins, which in turn enhanced VM formation in GBM. In summary, we clarified a novel pathway for RBMS3-induced circHECTD1 encoding functional peptide 463aa to mediate the ubiquitination of NR2F1, which inhibited VM formation in GBM. This study aimed to reveal new mechanisms of GBM progression in order to provide novel approaches and strategies for the anti-vascular therapy of GBM. The schematic illustration showed the inhibitory effect of circHECTD1-463aa in the VM formation in GBM.
Assuntos
Glioblastoma , Humanos , Glioblastoma/genética , Glioblastoma/patologia , Linhagem Celular Tumoral , Cromatografia Líquida , Espectrometria de Massas em Tandem , Peptídeos/genética , Peptídeos/metabolismo , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Transativadores/metabolismo , Proteínas de Ligação a RNARESUMO
Vasculogenic mimicry (VM) is an endothelium-independent tumor microcirculation that provides adequate blood supply for tumor growth. The presence of VM greatly hinders the treatment of glioblastoma (GBM) with anti-angiogenic drugs. Therefore, targeting VM formation may be a feasible therapeutic strategy for GBM. The research aimed to evaluate the roles of BUD13, CDK12, MBNL1 in regulating VM formation of GBM. BUD13 and CDK12 were upregulated and MBNL1 was downregulated in GBM tissues and cells. Knockdown of BUD13, CDK12, or overexpression of MBNL1 inhibited GBM VM formation. METTL3 enhanced the stability of BUD13 mRNA and upregulated its expression through m6A methylation. BUD13 enhanced the stability of CDK12 mRNA and upregulated its expression. CDK12 phosphorylated MBNL1, thereby regulating VM formation of GBM. The simultaneous knockdown of BUD13, CDK12, and overexpression of MBNL1 reduced the volume of subcutaneously transplanted tumors in nude mice and prolonged the survival period. Thus, the BUD13/CDK12/MBNL1 axis plays a crucial role in regulating VM formation of GBM and provides a potential target for GBM therapy.
Assuntos
Quinases Ciclina-Dependentes , Proteínas de Ligação a DNA , Glioblastoma , Metiltransferases , Proteínas de Ligação a RNA , Animais , Camundongos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Glioblastoma/irrigação sanguínea , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , Metilação , Camundongos Nus , Fosforilação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Microcirculação/genética , Microcirculação/fisiologia , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismoRESUMO
The blood-tumor barrier (BTB) contributes to poor therapeutic efficacy by limiting drug uptake; therefore, elevating BTB permeability is essential for glioma treatment. Here, we prepared astrocyte microvascular endothelial cells (ECs) and glioma microvascular ECs (GECs) as in vitro blood-brain barrier (BBB) and BTB models. Upregulation of METTL3 and IGF2BP3 in GECs increased the stability of CPEB2 mRNA through its m6A methylation. CPEB2 bound to and increased SRSF5 mRNA stability, which promoted the ETS1 exon inclusion. P51-ETS1 promoted the expression of ZO-1, occludin, and claudin-5 transcriptionally, thus regulating BTB permeability. Subsequent in vivo knockdown of these molecules in glioblastoma xenograft mice elevated BTB permeability, promoted doxorubicin penetration, and improved glioma-specific chemotherapeutic effects. These results provide a theoretical and experimental basis for epigenetic regulation of the BTB, as well as insight into comprehensive glioma treatment.
Assuntos
Neoplasias Encefálicas , Glioma , Metiltransferases , Proteínas de Ligação a RNA , Fatores de Processamento de Serina-Arginina , Animais , Astrócitos/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Células Endoteliais/metabolismo , Epigênese Genética , Glioma/tratamento farmacológico , Glioma/genética , Glioma/metabolismo , Humanos , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Microvasos/metabolismo , Permeabilidade , Fatores de Processamento de RNA/genética , Fatores de Processamento de RNA/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Fatores de Processamento de Serina-Arginina/genética , Fatores de Processamento de Serina-Arginina/metabolismo , Proteína da Zônula de Oclusão-1/genética , Proteína da Zônula de Oclusão-1/metabolismoRESUMO
BACKGROUND: RNA-binding proteins (RBPs) and circular RNAs (circRNAs) play important roles in glioblastoma multiforme (GBM). Aerobic glycolysis is a metabolic characteristic of GBM. However, the roles of RBPs and circRNAs in aerobic glycolysis in GBM remain unclear. The aim of this study is to explore the mechanisms by which RBPs and circRNAs regulate aerobic glycolysis in GBM cells. METHODS: RNA sequencing and circRNA microarray analysis were performed to identify RBPs and circRNAs for further study. Mass spectrometry validated the encoded protein and its interacting proteins. Quantitative reverse transcription PCR and western blot assays were used to determine the mRNA and protein expression, respectively. Furthermore, immunofluorescence and fluorescence in situ hybridization assays were used to determine the protein and RNA localization, respectively. Glucose and lactate measurement assays, Seahorse XF glycolysis stress assays and cell viability assays were conducted to investigate the effects on glycolysis and proliferation in GBM cells. RESULTS: We selected zinc finger CCHC-type and RNA-binding motif 1 (ZCRB1) and circRNA HEAT repeat containing 5B (circHEATR5B) as candidates for this study. These genes were expressed at low levels in GBM tissues and cells. Both ZCRB1 and circHEATR5B overexpression suppressed aerobic glycolysis and proliferation in GBM cells. ZCRB1 overexpression promoted the Alu element-mediated formation of circHEATR5B. In addition, circHEATR5B encoded a novel protein HEATR5B-881aa which interacted directly with Jumonji C-domain-containing 5 (JMJD5) and reduced its stability by phosphorylating S361. JMJD5 knockdown increased pyruvate kinase M2 (PKM2) enzymatic activity and suppressed glycolysis and proliferation in GBM cells. Finally, ZCRB1, circHEATR5B and HEATR5B-881aa overexpression inhibited GBM xenograft growth and prolonged the survival time of nude mice. CONCLUSIONS: This study reveals a novel mechanism of regulating aerobic glycolysis and proliferation in GBM cells through the ZCRB1/circHEATR5B/HEATR5B-881aa/JMJD5/PKM2 pathway, which can provide novel strategies and potential targets for GBM therapy.