RESUMO
By establishing multi-omics pipelines, we uncover overexpression and gene copy-number alterations of nucleoporin-93 (NUP93), a nuclear pore component, in aggressive human mammary tumors. NUP93 overexpression enhances transendothelial migration and matrix invasion in vitro, along with tumor growth and metastasis in animal models. These findings are supported by analyses of two sets of naturally occurring mutations: rare oncogenic mutations and inactivating familial nephrotic syndrome mutations. Mechanistically, NUP93 binds with importins, boosts nuclear transport of importins' cargoes, such as ß-catenin, and activates MYC. Likewise, NUP93 overexpression enhances the ultimate nuclear transport step shared by additional signaling pathways, including TGF-ß/SMAD and EGF/ERK. The emerging addiction to nuclear transport exposes vulnerabilities of NUP93-overexpressing tumors. Congruently, myristoylated peptides corresponding to the nuclear translocation signals of SMAD and ERK can inhibit tumor growth and metastasis. Our study sheds light on an emerging hallmark of advanced tumors, which derive benefit from robust nucleocytoplasmic transport.
Assuntos
Neoplasias da Mama , Complexo de Proteínas Formadoras de Poros Nucleares , Transporte Ativo do Núcleo Celular , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Feminino , Humanos , Poro Nuclear/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Circular RNAs (circRNAs) are a class of endogenous RNAs characterized by a covalent loop structure. In comparison to other types of RNAs, the abundance of circRNAs is relatively low but due to the circular configuration, their stability is very high. In addition, circRNAs display high degree of tissue specificity. The sponging activity of circRNAs toward microRNAs is the best-described mode of action of circRNAs. However, the ability of circRNAs to bind with specific proteins, as well as to encode short proteins, propose alternative functions. This review introduces the biogenesis of circRNAs and summarizes the roles played by circRNAs in human diseases. These include examples of their functional roles in several organ-specific cancers, such as head and neck and breast and lung cancers. In addition, we review potential functions of circRNAs in diabetes, cardiovascular, and neurodegenerative diseases. Recently, a growing number of studies have demonstrated involvement of circRNAs in a wide spectrum of signaling molecular pathways, but at the same time many different and controversial views on circRNAs role and function are emerging. We conclude by offering cellular homeostasis generated by networks comprising circular RNAs, other non-coding RNAs and RNA-binding proteins. Accordingly, it is predictable that circRNAs, due to their highly stable nature and remarkable tissue specificity, will emerge as reliable biomarkers of disease course and treatment efficacy.
Assuntos
Biomarcadores Tumorais/metabolismo , Doença/genética , RNA Circular/genética , HumanosRESUMO
HER2 upregulation is related with poor outcome in many tumor types. Whereas anti-HER2 treatment is the standard approach as adjuvant therapy in HER2-overexpressing breast cancer, the frequent relapses reinforce the need for alternative treatments. Here we used next-generation sequencing (NGS) to evaluate miRNAs and circRNAs in the cell-lines HB4a and C5.2, where the latter is a HER2-overexpressing clone of the former, and also from two different populations of their secreted extracellular vesicles. Whereas circRNA-levels were stable, we found at least 16 miRNAs apparently modulated by HER2-expression. The miR223-3p, miR-421 and miR-21-5p were validated in an independent cohort of 431 breast cancer patients from The Cancer Genome Atlas (TCGA). The consistent modulation of these molecules and their possible involvement in the HER2-axis makes them promising new targets to overcome HER2-activation.
Assuntos
Neoplasias da Mama/genética , MicroRNAs/genética , RNA Circular/genética , Receptor ErbB-2/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Transcriptoma/genéticaRESUMO
Activation of brown adipose tissue (BAT) controls energy homeostasis in rodents and humans and has emerged as an innovative strategy for the treatment of obesity and type 2 diabetes mellitus. Here we show that ageing- and obesity-associated dysfunction of brown fat coincides with global microRNA downregulation due to reduced expression of the microRNA-processing node Dicer1. Consequently, heterozygosity of Dicer1 in BAT aggravated diet-induced-obesity (DIO)-evoked deterioration of glucose metabolism. Analyses of differential microRNA expression during preadipocyte commitment and mouse models of progeria, longevity and DIO identified miR-328 as a regulator of BAT differentiation. Reducing miR-328 blocked preadipocyte commitment, whereas miR-328 overexpression instigated BAT differentiation and impaired muscle progenitor commitment-partly through silencing of the ß-secretase Bace1. Loss of Bace1 enhanced brown preadipocyte specification in vitro and was overexpressed in BAT of obese and progeroid mice. In vivo Bace1 inhibition delayed DIO-induced weight gain and improved glucose tolerance and insulin sensitivity. These experiments reveal Dicer1-miR-328-Bace1 signalling as a determinant of BAT function, and highlight the potential of Bace1 inhibition as a therapeutic approach to improve not only neurodegenerative diseases but also ageing- and obesity-associated impairments of BAT function.
Assuntos
Tecido Adiposo Marrom/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Ácido Aspártico Endopeptidases/genética , Diferenciação Celular/fisiologia , RNA Helicases DEAD-box/genética , MicroRNAs/genética , Ribonuclease III/genética , Secretases da Proteína Precursora do Amiloide/metabolismo , Animais , Ácido Aspártico Endopeptidases/metabolismo , RNA Helicases DEAD-box/metabolismo , Metabolismo Energético/fisiologia , Homeostase/fisiologia , Resistência à Insulina/fisiologia , Camundongos Endogâmicos C57BL , MicroRNAs/metabolismo , Obesidade/genética , Obesidade/metabolismo , Ribonuclease III/metabolismoRESUMO
ERBB2/HER2 é um gene frequentemente amplificado em cânceres de mama, estômago e ovário e está diretamente associado ao mal prognóstico de pacientes portadores desta alteração genética. A proteína HER2, super-expressa nestes tumores, é um importante alvo de terapias dirigidas e tem um papel relevante no tratamento de tumores com esta característica molecular. Entretanto, o surgimento de resistência aos tratamentos ainda é um desafio a ser superado. Em 2011, foi identificado o miR-4728-3p dentro de uma região intrônica de ERBB2/HER2, cuja super-expressão acompanhava a amplificação do oncogene. Pouco ainda se sabe sobre a função e possíveis alvos deste microRNA, o que motivou os estudos desta tese, onde usamos técnicas como RNA-Seq e proteômica buscando a identificação de alvos, seguidos de comparação com bancos de dados, além de ensaios in vitro e in vivo. Genes como RNASEH1, PSMC3, TUFM e GNA11 mostraram-se fortes candidatos-alvo do miR-4728-3p, sendo regulados após super-expressão e knockdown do microRNA em diferentes linhagens celulares. O bloqueio de miR-4728-3p em tumores gástricos HER2+ levou à diminuição do volume e peso tumoral, sugerindo um importante papel terapêutico no controle de tumores com amplificação de ERBB2/HER2. Os resultados obtidos indicam uma possível participação do miR-4728-3p na carcinogênese de tumores gástricos HER2+ e ampliam o conhecimento sobre a biologia de ERBB2/HER2. A identificação de alvos deste microRNA pode adicionalmente elucidar os mecanismos pelo qual o bloqueio do miR-4728-3p interfere no crescimento tumoral, levantando informações importantes sobre o controle da expressão gênica mediada por RNAs não- codificantes no câncer
The ERBB2/HER2 gene is frequently amplified in breast, stomach and ovarian cancers and is directly associated with poor prognosis. The HER2 protein, overexpressed in these tumors, is an important therapeutic target and plays relevant roles in tumor biology and prognosis. However, the emergence of resistance to anti-HER2 treatment is still a challenge to be overcomed. In 2011, miR-4728-3p was identified within an intronic region of the ERBB2/HER2 gene; when this region is amplified, both transcripts are also amplified and overexpressed. Little is known about the function and possible targets of this microRNA, which motivated the studies of this thesis, where techniques such as RNA-Seq and proteomics were employed to identify targets, followed by database analyses, in vitro and in vivo assays. Genes such as RNASEH1, PSMC3, TUFM and GNA11 were found to be strong target-candidates of miR-4728-3p and were regulated after the overexpression or knockdown of this miRNA in different cell lines. miR-4728-3p blockade in HER2+ gastric tumors of mouse xenografts led to decreased tumor volume and weight, suggesting its important therapeutic potential in the control of ERBB2/HER2 amplified tumors. The results obtained here indicate a possible participation of miR-4728-3p in the carcinogenesis of HER2+ gastric tumors and expand the knowledge of ERBB2/HER2 biology. The identification of targets of this microRNA may further elucidate the mechanisms by which miR-4728-3p blockade interferes with tumor growth, raising important information about the control of gene expression mediated by noncoding RNAs in cancer