RESUMO
KDM4 histone demethylases mainly catalyze the removal of methyl marks from H3K9 and H3K36 to epigenetically regulate chromatin structure and gene expression. KDM4 expression is strictly regulated to ensure proper function in a myriad of biological processes, including transcription, cellular proliferation and differentiation, DNA damage repair, immune response, and stem cell self-renewal. Aberrant expression of KDM4 demethylase has been documented in many types of blood and solid tumors, and thus, KDM4s represent promising therapeutic targets. In this chapter, we summarize the current knowledge of the structures and regulatory mechanisms of KDM4 proteins and our understanding of their alterations in human pathological processes with a focus on development and cancer. We also review the reported KDM4 inhibitors and discuss their potential as therapeutic agents.
Assuntos
Histona Desmetilases com o Domínio Jumonji , Neoplasias , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases com o Domínio Jumonji/química , Histona Desmetilases com o Domínio Jumonji/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Reparo do DNA , Proliferação de Células , Diferenciação Celular , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Histona Desmetilases/uso terapêutico , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêuticoRESUMO
Lysine acetyltransferases (KATs) are a highly diverse group of epigenetic enzymes that play important roles in various cellular processes including transcription, signal transduction, and cellular metabolism. However, our knowledge of the genomic and transcriptomic alterations of KAT genes and their clinical significance in human cancer remains incomplete. We undertook a metagenomic analysis of 37 KATs in more than 10 000 cancer samples across 33 tumor types, focusing on breast cancer. We identified associations among recurrent genetic alteration, gene expression, clinicopathologic features, and patient survival. Loss-of-function analysis was carried out to examine which KAT has important roles in growth and viability of breast cancer cells. We identified that a subset of KAT genes, including NAA10, KAT6A, and CREBBP, have high frequencies of genomic amplification or mutation in a spectrum of human cancers. Importantly, we found that 3 KATs, NAA10, ACAT2, and BRD4, were highly expressed in the aggressive basal-like subtype, and their expression was significantly associated with disease-free survival. Furthermore, we showed that depletion of NAA10 inhibits basal-like breast cancer growth in vitro. Our findings provide a strong foundation for further mechanistic research and for developing therapies that target NAA10 or other KATs in human cancer.
Assuntos
Genoma Humano/genética , Lisina Acetiltransferases/genética , Neoplasias/genética , Neoplasias/patologia , Neoplasias da Mama/classificação , Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Proteína de Ligação a CREB/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Intervalo Livre de Doença , Proteína p300 Associada a E1A/genética , Dosagem de Genes , Expressão Gênica , Histona Acetiltransferases/genética , Humanos , Lisina Acetiltransferases/metabolismo , Mutação , Acetiltransferase N-Terminal A/genética , Acetiltransferase N-Terminal A/metabolismo , Acetiltransferase N-Terminal E/genética , Acetiltransferase N-Terminal E/metabolismo , Neoplasias/mortalidade , Prognóstico , Fatores Associados à Proteína de Ligação a TATA/genética , Fator de Transcrição TFIID/genética , Fatores de Transcrição/genéticaRESUMO
Autophagy, a lysosomal degradative pathway in response to nutrient limitation, plays an important regulatory role in lipid homeostasis upon energy demands. Here, we demonstrated that the endoplasmic reticulum-tethered, stress-sensing transcription factor cAMP-responsive element-binding protein, hepatic-specific (CREBH) functions as a major transcriptional regulator of hepatic autophagy and lysosomal biogenesis in response to nutritional or circadian signals. CREBH deficiency led to decreased hepatic autophagic activities and increased hepatic lipid accumulation upon starvation. Under unfed or during energy-demanding phases of the circadian cycle, CREBH is activated to drive expression of the genes encoding the key enzymes or regulators in autophagosome formation or autophagic process, including microtubule-associated protein 1B-light chain 3, autophagy-related protein (ATG)7, ATG2b, and autophagosome formation Unc-51 like kinase 1, and the genes encoding functions in lysosomal biogenesis and homeostasis. Upon nutrient starvation, CREBH regulates and interacts with peroxisome proliferator-activated receptor α (PPARα) and PPARγ coactivator 1α to synergistically drive expression of the key autophagy genes and transcription factor EB, a master regulator of lysosomal biogenesis. Furthermore, CREBH regulates rhythmic expression of the key autophagy genes in the liver in a circadian-dependent manner. In summary, we identified CREBH as a key transcriptional regulator of hepatic autophagy and lysosomal biogenesis for the purpose of maintaining hepatic lipid homeostasis under nutritional stress or circadian oscillation.-Kim, H., Williams, D., Qiu, Y., Song, Z., Yang, Z., Kimler, V., Goldberg, A., Zhang, R., Yang, Z., Chen, X., Wang, L., Fang, D., Lin, J. D., Zhang, K. Regulation of hepatic autophagy by stress-sensing transcription factor CREBH.
Assuntos
Autofagia/fisiologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Privação de Alimentos/fisiologia , Regulação da Expressão Gênica/fisiologia , Fígado/metabolismo , Animais , Autofagossomos/metabolismo , Autofagia/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Ritmo Circadiano , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/deficiência , Fígado Gorduroso/etiologia , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Hepatócitos/metabolismo , Metabolismo dos Lipídeos , Fígado/citologia , Lisossomos/metabolismo , Camundongos , Camundongos Knockout , PPAR alfa/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Estresse Fisiológico/genética , Estresse Fisiológico/fisiologia , Transcrição GênicaRESUMO
RNA methylation, catalysed by a set of RNA methyltransferases (RNMTs), modulates RNA structures, properties, and biological functions. RNMTs are increasingly documented to be dysregulated in various human diseases, particularly developmental disorders and cancer. However, the genomic and transcriptomic alterations of RNMTs, as well as their functional roles in human cancer, are limited. In this study, we utilized an unbiased approach to examine copy number alterations and mutation rates of 58 RNMTs in more than 10,000 clinical samples across 32 human cancer types. We also investigated these alterations and RNMT expression level as they related to clinical features such as tumour subtype, grade, and survival in a large cohort of tumour samples, focusing on breast cancer. Loss-of-function analysis was performed to examine RNMT candidates with important roles in growth and viability of breast cancer cells. We identified a subset of RNMTs, notably TRMT12, NSUN2, TARBP1, and FTSJ3, that were amplified or mutated in a subset of human cancers. Several RNMTs were significantly associated with breast cancer aggressiveness and poor prognosis. Loss-of-function analysis indicated FTSJ3, a 2'-O-Me methyltransferase, as a candidate RNMT with functional roles in promoting cancer growth and survival. A subset of RNMTs, like FTSJ3, represents promising novel targets for anticancer drug discovery. Our findings provide a framework for further study of the functional consequences of RNMT alterations in human cancer and for developing therapies that target cancer-promoting RNMTs in the future.
Assuntos
Neoplasias da Mama/genética , Mutação com Perda de Função , Metiltransferases/genética , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Progressão da Doença , Feminino , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , PrognósticoRESUMO
BACKGROUND: Autoantibodies function as markers of tumorigenesis and have been proposed to enhance early detection of malignancies. We recently reported, using immunoscreening of a T7 complementary DNA (cDNA) library of breast cancer (BC) proteins with sera from patients with BC, the presence of autoantibodies targeting several mitochondrial DNA (mtDNA)-encoded subunits of the electron transport chain (ETC) in complexes I, IV, and V. METHODS: In this study, we have characterized the role of Mitochondrial-Nuclear Retrograde Regulator 1 (MNRR1, also known as CHCHD2), identified on immunoscreening, in breast carcinogenesis. We assessed the protein as well as transcript levels of MNRR1 in BC tissues and in derived cell lines representing tumors of graded aggressiveness. Mitochondrial function was also assayed and correlated with the levels of MNRR1. We studied the invasiveness of BC derived cells and the effect of MNRR1 levels on expression of genes associated with cell proliferation and migration such as Rictor and PGC-1α. Finally, we manipulated levels of MNRR1 to assess its effect on mitochondria and on some properties linked to a metastatic phenotype. RESULTS: We identified a nuclear DNA (nDNA)-encoded mitochondrial protein, MNRR1, that was significantly associated with the diagnosis of invasive ductal carcinoma (IDC) of the breast by autoantigen microarray analysis. In focusing on the mechanism of action of MNRR1 we found that its level was nearly twice as high in malignant versus benign breast tissue and up to 18 times as high in BC cell lines compared to MCF10A control cells, suggesting a relationship to aggressive potential. Furthermore, MNRR1 affected levels of multiple genes previously associated with cancer metastasis. CONCLUSIONS: MNRR1 regulates multiple genes that function in cell migration and cancer metastasis and is higher in cell lines derived from aggressive tumors. Since MNRR1 was identified as an autoantigen in breast carcinogenesis, the present data support our proposal that both mitochondrial autoimmunity and MNRR1 activity in particular are involved in breast carcinogenesis. Virtually all other nuclear encoded genes identified on immunoscreening of invasive BC harbor an MNRR1 binding site in their promoters, thereby placing MNRR1 upstream and potentially making it a novel marker for BC metastasis.
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Neoplasias da Mama/diagnóstico , Carcinoma Ductal de Mama/diagnóstico , Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Autoantígenos/metabolismo , Autoimunidade , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Carcinoma Ductal de Mama/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Proteínas de Ligação a DNA , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Mitocôndrias/genética , Invasividade Neoplásica , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Estudos Prospectivos , Análise Serial de Proteínas , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Regulação para CimaRESUMO
Histone lysine demethylases (KDMs) comprise a large class of enzymes that catalyze site-specific demethylation of lysine residues on histones and other proteins. They play critical roles in controlling transcription, chromatin architecture, and cellular differentiation. However, the genomic landscape and clinical significance of KDMs in breast cancer remain poorly characterized. Here, we conducted a meta-analysis of 24 KDMs in breast cancer and identified associations among recurrent copy number alterations, gene expression, breast cancer subtypes, and clinical outcome. Two KDMs, KDM2A and KDM5B, had the highest frequency of genetic amplification and overexpression. Furthermore, among the 24 KDM genes, KDM2A had the highest correlation between copy number and mRNA expression, and high mRNA levels of KDM2A were significantly associated with shorter survival of breast cancer patients. KDM2A has two isoforms: the long isoform is comprised of a JmjC domain, CXXC-zinc finger, PHD zinc finger, F-box, and the AMN1 protein domain; whereas the short isoform of KDM2A lacks the N-terminal JmjC domain but contains all other motifs. Detailed characterization of KDM2A in breast cancer revealed that the short isoform of KDM2A is more abundant than the long isoform at DNA, mRNA, and protein levels in a subset of breast cancers. Furthermore, our data indicate that the short isoform of KDM2A has oncogenic potential and functions as an oncogenic isoform in a subset of breast cancers. Taken together, our findings suggest that amplification and overexpression of the KDM2A short isoform is critical in breast cancer progression.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Humanos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Análise de SobrevidaRESUMO
Basal-like breast cancer (BLBC) accounts for the most aggressive types of breast cancer, marked by high rates of relapse and poor prognoses and with no effective clinical therapy yet. Therefore, investigation of new targets and treatment strategies is more than necessary. Here, we identified a receptor that can be targeted in BLBC for efficient and specific siRNA mediated gene knockdown of therapeutically relevant genes such as the histone demethylase GASC1, which is involved in multiple signaling pathways leading to tumorigenesis. Breast cancer and healthy breast cell lines were compared regarding transferrin receptor (TfR) expression via flow cytometry and transferrin binding assays. Nanobioconjugates made of low molecular weight polyethylenimine (LMW-PEI) and transferrin (Tf) were synthesized to contain a bioreducible disulfide bond. siRNA complexation was characterized by condensation assays and dynamic light scattering. Cytotoxicity, transfection efficiency, and the targeting specificity of the conjugates were investigated in TfR positive and negative healthy breast and breast cancer cell lines by flow cytometry, confocal microscopy, RT-PCR, and Western blot. Breast cancer cell lines revealed a significantly higher TfR expression than healthy breast cells. The conjugates efficiently condensed siRNA into particles with 45 nm size at low polymer concentrations, showed no apparent toxicity on different breast cancer cell lines, and had significantly greater transfection and gene knockdown activity on mRNA and protein levels than PEI/siRNA leading to targeted and therapeutic growth inhibition post GASC1 knockdown. The synthesized nanobioconjugates improved the efficiency of gene transfer and targeting specificity in transferrin receptor positive cells but not in cells with basal receptor expression. Therefore, these materials in combination with our newly identified siRNA sequences are promising candidates for therapeutic targeting of hard-to-treat BLBC and are currently further investigated regarding in vivo targeting efficacy and biocompatibility.
Assuntos
Regulação da Expressão Gênica/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Nanopartículas/química , RNA Interferente Pequeno/genética , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/genética , Proliferação de Células/fisiologia , Feminino , Citometria de Fluxo , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Microscopia de Força Atômica , Microscopia Confocal , Polímeros/química , Receptores da Transferrina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transferrina/metabolismoRESUMO
Increased de novo lipogenesis is a hallmark of aggressive cancers. Lipid droplets, the major form of cytosolic lipid storage, have been implicated in cancer cell proliferation and tumorigenesis. Recently, we identified the ERLIN2 [ER (endoplasmic reticulum) lipid raft-associated 2) gene that is amplified and overexpressed in aggressive human breast cancer. Previous studies demonstrated that ERLIN2 plays a supporting oncogenic role by facilitating the transformation of human breast cancer cells. In the present study, we found that ERLIN2 supports cancer cell growth by regulating cytosolic lipid droplet production. ERLIN2 is preferably expressed in human breast cancer cells or hepatoma cells and is inducible by insulin signalling or when cells are cultured in lipoprotein-deficient medium. Increased expression of ERLIN2 promotes the accumulation of cytosolic lipid droplets in breast cancer cells or hepatoma cells in response to insulin or overload of unsaturated fatty acids. ERLIN2 regulates activation of SREBP (sterol regulatory element-binding protein) 1c, the key regulator of de novo lipogenesis, in cancer cells. ERLIN2 was found to bind to INSIG1 (insulin-induced gene 1), a key ER membrane protein that blocks SREBP activation. Consistent with the role of ERLIN2 in regulating cytosolic lipid content, down-regulation of ERLIN2 in breast cancer or hepatoma cells led to lower cell proliferation rates. The present study revealed a novel role for ERLIN2 in supporting cancer cell growth by promoting the activation of the key lipogenic regulator SREBP1c and the production of cytosolic lipid droplets. The identification of ERLIN2 as a regulator of cytosolic lipid content in cancer cells has important implications for understanding the molecular basis of tumorigenesis and the treatment of cancer.
Assuntos
Neoplasias da Mama/genética , Citosol/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/genética , Neoplasias da Mama/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipídeos/fisiologia , Proteínas de Membrana/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismoRESUMO
BACKGROUND: Amplification of the 8p11-12 region has been found in approximately 15% of human breast cancer and is associated with poor prognosis. Previous genomic analysis has led us to identify the endoplasmic reticulum (ER) lipid raft-associated 2 (ERLIN2) gene as one of the candidate oncogenes within the 8p11-12 amplicon in human breast cancer, particularly in the luminal subtype. ERLIN2, an ER membrane protein, has recently been identified as a novel mediator of ER-associated degradation. Yet, the biological roles of ERLIN2 and molecular mechanisms by which ERLIN2 coordinates ER pathways in breast carcinogenesis remain unclear. METHODS: We established the MCF10A-ERLIN2 cell line, which stably over expresses ERLIN2 in human nontransformed mammary epithelial cells (MCF10A) using the pLenti6/V5-ERLIN2 construct. ERLIN2 over expressing cells and their respective parental cell lines were assayed for in vitro transforming phenotypes. Next, we knocked down the ERLIN2 as well as the ER stress sensor IRE1α activity in the breast cancer cell lines to characterize the biological roles and molecular basis of the ERLIN2 in carcinogenesis. Finally, immunohistochemical staining was performed to detect ERLIN2 expression in normal and cancerous human breast tissues RESULTS: We found that amplification of the ERLIN2 gene and over expression of the ERLIN2 protein occurs in both luminal and Her2 subtypes of breast cancer. Gain- and loss-of-function approaches demonstrated that ERLIN2 is a novel oncogenic factor associated with the ER stress response pathway. The IRE1α/XBP1 axis in the ER stress pathway modulated expression of ERLIN2 protein levels in breast cancer cells. We also showed that over expression of ERLIN2 facilitated the adaptation of breast epithelial cells to ER stress by supporting cell growth and protecting the cells from ER stress-induced cell death. CONCLUSIONS: ERLIN2 may confer a selective growth advantage for breast cancer cells by facilitating a cytoprotective response to various cellular stresses associated with oncogenesis. The information provided here sheds new light on the mechanism of breast cancer malignancy.
Assuntos
Neoplasias da Mama/genética , Estresse do Retículo Endoplasmático/genética , Proteínas de Membrana/genética , Transdução de Sinais , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Análise por Conglomerados , Variações do Número de Cópias de DNA , Proteínas de Ligação a DNA/metabolismo , Endorribonucleases/metabolismo , Feminino , Amplificação de Genes , Expressão Gênica , Humanos , Glândulas Mamárias Humanas/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , Fatores de Transcrição de Fator Regulador X , Fatores de Transcrição/metabolismo , Proteína 1 de Ligação a X-BoxRESUMO
SARS-CoV-2, an emerging coronavirus, has spread rapidly around the world, resulting in over ten million cases and more than half a million deaths as of July 1, 2020. Effective treatments and vaccines for SARS-CoV-2 infection do not currently exist. Previous studies demonstrated that nonstructural protein 16 (nsp16) of coronavirus is an S-adenosyl methionine (SAM)-dependent 2'-O-methyltransferase (2'-O-MTase) that has an important role in viral replication and prevents recognition by the host innate immune system. In the present study, we employed structural analysis, virtual screening, and molecular simulation approaches to identify clinically investigated and approved drugs which can act as promising inhibitors against nsp16 2'-O-MTase of SARS-CoV-2. Comparative analysis of primary amino acid sequences and crystal structures of seven human CoVs defined the key residues for nsp16 2-O'-MTase functions. Virtual screening and docking analysis ranked the potential inhibitors of nsp16 from more than 4,500 clinically investigated and approved drugs. Furthermore, molecular dynamics simulations were carried out on eight top candidates, including Hesperidin, Rimegepant, Gs-9667, and Sonedenoson, to calculate various structural parameters and understand the dynamic behavior of the drug-protein complexes. Our studies provided the foundation to further test and repurpose these candidate drugs experimentally and/or clinically for COVID-19 treatment.Communicated by Ramaswamy H. Sarma.
Assuntos
Tratamento Farmacológico da COVID-19 , SARS-CoV-2 , Vacinas contra COVID-19 , Humanos , Metiltransferases , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , RiboseRESUMO
Human methyltransferase-like (METTL) proteins transfer methyl groups to nucleic acids, proteins, lipids, and other small molecules, subsequently playing important roles in various cellular processes. In this study, we performed integrated genomic, transcriptomic, proteomic, and clinicopathological analyses of 34 METTLs in a large cohort of primary tumor and cell line data. We identified a subset of METTL genes, notably METTL1, METTL7B, and NTMT1, with high frequencies of genomic amplification and/or up-regulation at both the mRNA and protein levels in a spectrum of human cancers. Higher METTL1 expression was associated with high-grade tumors and poor disease prognosis. Loss-of-function analysis in tumor cell lines indicated the biological importance of METTL1, an m7G methyltransferase, in cancer cell growth and survival. Furthermore, functional annotation and pathway analysis of METTL1-associated proteins revealed that, in addition to the METTL1 cofactor WDR4, RNA regulators and DNA packaging complexes may be functionally interconnected with METTL1 in human cancer. Finally, we generated a crystal structure model of the METTL1-WDR4 heterodimeric complex that might aid in understanding the key functional residues. Our results provide new information for further functional study of some METTL alterations in human cancer and might lead to the development of small inhibitors that target cancer-promoting METTLs.
Assuntos
Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Neoplasias/patologia , Linhagem Celular Tumoral , Proliferação de Células , Proteínas de Ligação ao GTP/química , Amplificação de Genes , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genômica , Humanos , Metiltransferases/química , Modelos Moleculares , Gradação de Tumores , Neoplasias/genética , Neoplasias/metabolismo , Prognóstico , Ligação Proteica , Conformação Proteica , Proteômica , Análise de SobrevidaRESUMO
Dysregulation of inositol-requiring enzyme 1 (IRE1), the primary transducer of Unfolded Protein Response (UPR), has been observed in tumor initiation and progression, but the underlying mechanism remains to be further elucidated. In this study, we identified that the IRE1 gene is frequently amplified and over-expressed in aggressive luminal B breast cancer cells and that IRE1 upregulation is significantly associated with worse overall survival of patients with breast cancer. IRE1 processes and mediates degradation of a subset of tumor suppressor microRNAs (miRNAs), including miR-3607, miR-374a, and miR-96, via a mechanism called Regulated IRE1-Dependent Decay (RIDD). IRE1-dependent degradation of tumor suppressor miR-3607 leads to elevation of RAS oncogene GTPase RAB3B in breast cancer cells. Inhibition of IRE1 endoribonuclease activity with the pharmacological compound 4µ8C or genetic approaches effectively suppresses luminal breast cancer cell proliferation and aggressive cancer phenotypes. Our work revealed the IRE1-RIDD-miRNAs pathway that promotes malignancy of luminal breast cancer.
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Recently, we analysed the 8p11-12 genomic region for copy number and gene expression changes in a panel of human breast cancer cell lines and primary specimens. We found that SFRP1 (Secreted frizzled related protein 1) is frequently under expressed even in breast tumours with copy number increases in this genomic region. SFRP1 encodes a WNT signalling antagonist, and plays a role in the development of multiple solid tumour types. In this study, we analysed methylation-associated silencing of the SFRP1 gene in breast cancer cells with the 8p11-12 amplicon, and investigated the tumour suppressor properties of SFRP1 in breast cancer cells. SFRP1 expression was markedly reduced in both the breast cancer cell lines and primary tumour specimens relative to normal primary human mammary epithelial cells even when SFRP1 is amplified. Suppression of SFRP1 expression in breast cancer cells with an SFRP1 gene amplification is associated with SFRP1 promoter methylation. Furthermore, restoration of SFRP1 expression suppressed the growth of breast cancer cells in monolayer, and inhibited anchorage independent growth. We also examined the relationship between the silencing of SFRP1 gene and WNT signalling in breast cancer. Ectopic SFRP1 expression in breast cancer cells suppressed both canonical and non-canonical WNT signalling pathways, and SFRP1 expression was negatively associated with the expression of a subset of WNT responsive genes including RET and MSX2. Thus, down-regulation of SFRP1 can be triggered by epigenetic and/or genetic events and may contribute to the tumourigenesis of human breast cancer through both canonical and non-canonical WNT signalling pathways.
Assuntos
Neoplasias da Mama/genética , Metilação de DNA , Inativação Gênica , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de Membrana/genética , Proteínas Wnt/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/genética , Humanos , Proteínas Proto-Oncogênicas c-ret/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética , Transdução Genética , TransfecçãoRESUMO
More than 20 different PIK3CA gene mutations were identified in breast cancer with different frequencies. Whether these breast cancer associated mutations have similar biological effects is largely unknown. In this study, we established a novel cell model using the lentivirus system to express 10 different PIK3CA genes (wild type and mutant) based on the human mammary epithelial cell MCF10A. We found that nine different PIK3CA mutants harbor different abilities to promote cell proliferation and EGF independent growth. In addition, most PIK3CA mutants (except for the wild type PIK3CA, the Q60K and the K111N mutants) had the ability to change the morphogenesis of the MCF10A cell in 3D Matrigel assay. Moreover, different PIK3CA mutants have different abilities to promote colony formation and cell invasion. We further observed that most of the PIK3CA mutants could activate p-AKT and p-p70-S6K in the absence of EGF stimulation. Finally, LY294002, a PI3K inhibitor, can effectively inhibit cell growth in cell lines with different PIK3CAs. Taken together, our results support the notion that different PIK3CA mutations differentially contribute to breast cancer transformation, and exploration of the therapeutic application of these mutations will benefit breast cancer patients with the PIK3CA mutations.
Assuntos
Neoplasias da Mama/patologia , Células Epiteliais/metabolismo , Glândulas Mamárias Humanas/metabolismo , Fosfatidilinositol 3-Quinases/genética , Linhagem Celular Tumoral , Proliferação de Células , Cromonas/farmacologia , Classe I de Fosfatidilinositol 3-Quinases , Colágeno/química , Combinação de Medicamentos , Inibidores Enzimáticos/farmacologia , Fator de Crescimento Epidérmico/metabolismo , Humanos , Laminina/química , Lentivirus/metabolismo , Glândulas Mamárias Humanas/patologia , Morfolinas/farmacologia , Mutação , Fosfatidilinositol 3-Quinases/metabolismo , Proteoglicanas/química , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismoRESUMO
We previously reported that expression of an environmentally induced gene, mineral dust-induced gene (mdig), predicts overall survival in breast cancer patients. In the present report, we further demonstrate the differential roles of mdig between earlier- and later-stage breast cancers. In noncancerous breast, mdig is a proliferation factor for cell growth and cell motility. In breast cancer, however, higher levels of mdig negatively regulate the migration and invasion of cancer cells. Assessment of global DNA methylation, chromatin accessibility and H3K9me3 heterochromatin signature suggests that silencing mdig enhances DNA and histone methylation. Through immunostaining and data mining, we found that mdig is significantly upregulated in noninvasive and/or earlier-stage breast cancers. In contrast, in triple-negative and other invasive breast cancers, diminished mdig expression was noted, indicating that the loss of mdig expression could be an important feature of aggressive breast cancers. Taken together, our data suggest that mdig is a new biomarker that likely promotes tumor growth in the early stages of breast cancer while acting as a tumor suppressor to inhibit invasion and metastasis in later-stage tumors.
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Diabetic skin ulcers represent a challenging clinical problem with mechanisms not fully understood. In this study, we investigated the role and mechanism for the primary unfolded protein response (UPR) transducer inositol-requiring enzyme 1 (IRE1α) in diabetic wound healing. Bone marrow-derived progenitor cells (BMPCs) were isolated from adult male type 2 diabetic and their littermate control mice. In diabetic BMPCs, IRE1α protein expression and phosphorylation were repressed. The impaired diabetic BMPC angiogenic function was rescued by adenovirus-mediated expression of IRE1α but not by the RNase-inactive IRE1α or the activated X-box binding protein 1 (XBP1), the canonical IRE1α target. In fact, IRE1α RNase processes a subset of microRNAs (miRs), including miR-466 and miR-200 families, through which IRE1α plays an important role in maintaining BMPC function under the diabetic condition. IRE1α attenuated maturation of miR-466 and miR-200 family members at precursor miR levels through the regulated IRE1α-dependent decay (RIDD) independent of XBP1. IRE1α deficiency in diabetes resulted in a burst of functional miRs from miR-466 and miR-200 families, which directly target and repress the mRNA encoding the angiogenic factor angiopoietin 1 (ANGPT1), leading to decreased ANGPT1 expression and disrupted angiogenesis. Importantly, cell therapies using IRE1α-expressing BMPCs or direct IRE1α gene transfer significantly accelerated cutaneous wound healing in diabetic mice through facilitating angiogenesis. In conclusion, our studies revealed a novel mechanistic basis for rescuing angiogenesis and tissue repair in diabetic wound treatments.
Assuntos
Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Proteínas de Membrana/metabolismo , MicroRNAs/genética , Proteínas Serina-Treonina Quinases/metabolismo , Regiões 3' não Traduzidas/genética , Angiopoietina-1/genética , Angiopoietina-1/metabolismo , Animais , Western Blotting , Terapia Baseada em Transplante de Células e Tecidos , Diabetes Mellitus Experimental/terapia , Eletroforese em Gel de Poliacrilamida , Feminino , Masculino , Proteínas de Membrana/genética , Camundongos , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Reação em Cadeia da Polimerase em Tempo Real , Cicatrização/genética , Cicatrização/fisiologia , Proteína 1 de Ligação a X-Box/genética , Proteína 1 de Ligação a X-Box/metabolismoRESUMO
Chromodomain helicase DNA binding proteins (CHDs) are characterized by N-terminal tandem chromodomains and a central adenosine triphosphate-dependent helicase domain. CHDs govern the cellular machinery's access to DNA, thereby playing critical roles in various cellular processes including transcription, proliferation, and DNA damage repair. Accumulating evidence demonstrates that mutation and dysregulation of CHDs are implicated in the pathogenesis of developmental disorders and cancer. However, we know little about genomic and transcriptomic alterations and the clinical significance of most CHDs in human cancer. We used TCGA and METABRIC datasets to perform integrated genomic and transcriptomic analyses of nine CHD genes in more than 10 000 primary cancer specimens from 32 tumor types, focusing on breast cancers. We identified associations among recurrent copy number alteration, gene expression, clinicopathological features, and patient survival. We found that CHD7 was the most commonly gained/amplified and mutated, whereas CHD3 was the most deleted across the majority of tumor types, including breast cancer. Overexpression of CHD7 was more prevalent in aggressive subtypes of breast cancer and was significantly correlated with high tumor grade and poor prognosis. CHD7 is required to maintain open, accessible chromatin, thus providing fine-tuning of transcriptional regulation of certain classes of genes. We found that CHD7 expression was positively correlated with a small subset of classical oncogenes, notably NRAS, in breast cancer. Knockdown of CHD7 inhibits cell proliferation and decreases gene expression of several CHD7 targets, including NRAS, in breast cancer cell lines. Thus, our results demonstrate the oncogenic potential of CHD7 and its association with poor prognostic parameters in human cancer.
Assuntos
DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Mutação , Neoplasias/genética , Transcriptoma , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Feminino , Dosagem de Genes , Genômica , Humanos , Masculino , Neoplasias/patologia , Oncogenes , Prognóstico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologiaRESUMO
A wide range of the epigenetic effectors that regulate chromatin modification, gene expression, genomic stability, and DNA repair contain structurally conserved domains called plant homeodomain (PHD) fingers. Alternations of several PHD finger-containing proteins (PHFs) due to genomic amplification, mutations, deletions, and translocations have been linked directly to various types of cancer. However, little is known about the genomic landscape and the clinical significance of PHFs in breast cancer. Hence, we performed a large-scale genomic and transcriptomic analysis of 98 PHF genes in breast cancer using TCGA and METABRIC datasets and correlated the recurrent alterations with clinicopathological features and survival of patients. Different subtypes of breast cancer had different patterns of copy number and expression for each PHF. We identified a subset of PHF genes that was recurrently altered with high prevalence, including PYGO2 (pygopus family PHD finger 2), ZMYND8 (zinc finger, MYND-type containing 8), ASXL1 (additional sex combs like 1) and CHD3 (chromodomain helicase DNA binding protein 3). Copy number increase and overexpression of ZMYND8 were more prevalent in Luminal B subtypes and were significantly associated with shorter survival of breast cancer patients. ZMYND8 was also involved in a positive feedback circuit of the estrogen receptor (ER) pathway, and the expression of ZMYND8 was repressed by the bromodomain and extra terminal (BET) inhibitor in breast cancer. Our findings suggest a promising avenue for future research-to focus on a subset of PHFs to better understand the molecular mechanisms and to identify therapeutic targets in breast cancer.
Assuntos
Neoplasias da Mama/genética , Variações do Número de Cópias de DNA , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Genômica/métodos , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Immunoblotting , Estimativa de Kaplan-Meier , Filogenia , Prognóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
TGF-ß-inducible early gene 1 (TIEG1), also known as Krüppel-like factor 10 (Klf10), represents a major downstream transcription factor of transforming growth factor-ß1 (TGF-ß1) signaling. Epidermal Langerhans cells (LCs), a unique subpopulation of dendritic cells (DC), essentially mediates immune surveillance and tolerance. TGF-ß1 plays a pivotal role in LC maintenance and function after birth, although the underpinning mechanisms remain elusive. Here, we hypothesized that TIEG1 might be involved in TGF-ß1-mediated LC homeostasis and function. Utilizing TIEG1 null mice, we discovered that TIEG1 deficiency did not alter LC homeostasis at the steady state and LC repopulation at inflamed-state, as well as their antigen-uptake capacity, but significantly impaired their maturation ability, which was opposite to the fact that loss of TGF-ß1 induced spontaneous LC maturation. Moreover, the ablation of TIEG1 enhanced skin contact hypersensitivity response. Our results suggested that TIEG1 is not a key molecule involved in TGF-ß1-mediated homeostasis, while TIEG1-related signaling pathways regulate LC maturation and their function.
RESUMO
There are 18 lysine deacetylases, also known as histone deacetylases (HDACs), that remove acetyl groups from histone and non-histone proteins, thereby playing critical roles in numerous biological processes. In many human cancers, HDACs are dysregulated through mutation, altered expression, or inappropriate recruitment to certain loci. However, knowledge of the genomic and transcriptomic alterations and the clinical significance of most HDACs in breast cancer remain incomplete. We used TCGA and METABRIC datasets to perform comprehensive, integrated genomic and transcriptomic analyses of 18 HDAC genes in approximately 3000 primary breast cancers and identified associations among recurrent copy number alteration, gene expression, clinicopathological features, and patient survival. We found distinct patterns of copy number alteration and expression for each HDAC in breast cancer subtypes. We demonstrated that HDAC2 and SIRT7 were the most commonly amplified/overexpressed, and SIRT3 was most deleted/underexpressed, particularly in aggressive basal-like breast cancer. Overexpression of HDAC2 was significantly correlated with high tumor grade, positive lymph node status, and poor prognosis. The HDAC inhibitor mocetinostat showed anti-tumor effects in HDAC2-overexpressing basal-like breast cancer lines in vitro. Furthermore, HDAC2 expression was positively correlated with a set of DNA-damage response genes, notably RAD51. We revealed a potential mechanism by which HDAC2 regulates RAD51 expression-by indirect mediation through microRNAs, e.g., miR-182. HDAC inhibitors have emerged as a promising new class of multifunctional anticancer agents. Identifying which breast cancers or patients show HDAC deregulation that contributes to tumor development/progression might enable us to improve target cancer therapy.