RESUMO
BACKGROUND: RNA editing generates modifications to the RNA sequences, thereby increasing protein diversity and shaping various layers of gene regulation. Recent studies have revealed global shifts in editing levels across many cancer types, as well as a few specific mechanisms implicating individual sites in tumorigenesis or metastasis. However, most tumor-associated sites, predominantly in noncoding regions, have unknown functional relevance. RESULTS: Here, we carry out integrative analysis of RNA editing profiles between epithelial and mesenchymal tumors, since epithelial-mesenchymal transition is a key paradigm for metastasis. We identify distinct editing patterns between epithelial and mesenchymal tumors in seven cancer types using TCGA data, an observation further supported by single-cell RNA sequencing data and ADAR perturbation experiments in cell culture. Through computational analyses and experimental validations, we show that differential editing sites between epithelial and mesenchymal phenotypes function by regulating mRNA abundance of their respective genes. Our analysis of RNA-binding proteins reveals ILF3 as a potential regulator of this process, supported by experimental validations. Consistent with the known roles of ILF3 in immune response, epithelial-mesenchymal differential editing sites are enriched in genes involved in immune and viral processes. The strongest target of editing-dependent ILF3 regulation is the transcript encoding PKR, a crucial player in immune and viral response. CONCLUSIONS: Our study reports widespread differences in RNA editing between epithelial and mesenchymal tumors and a novel mechanism of editing-dependent regulation of mRNA abundance. It reveals the broad impact of RNA editing in cancer and its relevance to cancer-related immune pathways.
Assuntos
Imunidade , Neoplasias/genética , Neoplasias/imunologia , Edição de RNA , RNA Mensageiro/genética , Células A549 , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Carcinogênese , Linhagem Celular Tumoral , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Proteínas do Fator Nuclear 90/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Análise de Sequência de RNARESUMO
Adenosine-to-inosine (A-to-I) editing, mediated by the ADAR enzymes, diversifies the transcriptome by altering RNA sequences. Recent studies reported global changes in RNA editing in disease and development. Such widespread editing variations necessitate an improved understanding of the regulatory mechanisms of RNA editing. Here, we study the roles of >200 RNA-binding proteins (RBPs) in mediating RNA editing in two human cell lines. Using RNA-sequencing and global protein-RNA binding data, we identify a number of RBPs as key regulators of A-to-I editing. These RBPs, such as TDP-43, DROSHA, NF45/90 and Ro60, mediate editing through various mechanisms including regulation of ADAR1 expression, interaction with ADAR1, and binding to Alu elements. We highlight that editing regulation by Ro60 is consistent with the global up-regulation of RNA editing in systemic lupus erythematosus. Additionally, most key editing regulators act in a cell type-specific manner. Together, our work provides insights for the regulatory mechanisms of RNA editing.
Assuntos
Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Regulação Neoplásica da Expressão Gênica , Edição de RNA/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Adenosina/genética , Elementos Alu , Autoantígenos/genética , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Inosina/genética , Células K562 , Lúpus Eritematoso Sistêmico/genética , RNA Citoplasmático Pequeno/genética , Ribonucleoproteínas/genética , Análise de Sequência de RNA , Transcrição Gênica , TransfecçãoRESUMO
Transcriptomic analyses of postmortem brains have begun to elucidate molecular abnormalities in autism spectrum disorder (ASD). However, a crucial pathway involved in synaptic development, RNA editing, has not yet been studied on a genome-wide scale. Here we profiled global patterns of adenosine-to-inosine (A-to-I) editing in a large cohort of postmortem brains of people with ASD. We observed a global bias for hypoediting in ASD brains, which was shared across brain regions and involved many synaptic genes. We show that the Fragile X proteins FMRP and FXR1P interact with RNA-editing enzymes (ADAR proteins) and modulate A-to-I editing. Furthermore, we observed convergent patterns of RNA-editing alterations in ASD and Fragile X syndrome, establishing this as a molecular link between these related diseases. Our findings, which are corroborated across multiple data sets, including dup15q (genomic duplication of 15q11.2-13.1) cases associated with intellectual disability, highlight RNA-editing dysregulation in ASD and reveal new mechanisms underlying this disorder.
Assuntos
Transtorno Autístico/metabolismo , Encéfalo/metabolismo , Edição de RNA , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Transtorno Autístico/genética , Proteína do X Frágil da Deficiência Intelectual/genética , Proteína do X Frágil da Deficiência Intelectual/metabolismo , Perfilação da Expressão Gênica , Humanos , Neurônios/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
Shelterin, a six-member complex, protects telomeres from nucleolytic attack and regulates their elongation by telomerase. Here, we have developed a strategy, called MICro-MS (Mapping Interfaces via Crosslinking-Mass Spectrometry), that combines crosslinking-mass spectrometry and phylogenetic analysis to identify contact sites within the complex. This strategy allowed identification of separation-of-function mutants of fission yeast Ccq1, Poz1, and Pot1 that selectively disrupt their respective interactions with Tpz1. The various telomere dysregulation phenotypes observed in these mutants further emphasize the critical regulatory roles of Tpz1-centered shelterin interactions in telomere homeostasis. Furthermore, the conservation between fission yeast Tpz1-Pot1 and human TPP1-POT1 interactions led us to map a human melanoma-associated POT1 mutation (A532P) to the TPP1-POT1 interface. Diminished TPP1-POT1 interaction caused by hPOT1-A532P may enable unregulated telomere extension, which, in turn, helps cancer cells to achieve replicative immortality. Therefore, our study reveals a connection between shelterin connectivity and tumorigenicity.
Assuntos
Aminopeptidases/metabolismo , Proteínas de Transporte/metabolismo , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Melanoma/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Serina Proteases/metabolismo , Neoplasias Cutâneas/metabolismo , Proteínas de Ligação a Telômeros/metabolismo , Aminopeptidases/química , Aminopeptidases/genética , Sítios de Ligação , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Proteínas de Transporte/química , Proteínas de Transporte/genética , Proteínas de Ligação a DNA , Dipeptidil Peptidases e Tripeptidil Peptidases/química , Dipeptidil Peptidases e Tripeptidil Peptidases/genética , Regulação Fúngica da Expressão Gênica , Humanos , Espectrometria de Massas/métodos , Melanoma/genética , Melanoma/patologia , Modelos Moleculares , Mutação , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/química , Proteínas de Schizosaccharomyces pombe/genética , Homologia de Sequência de Aminoácidos , Serina Proteases/química , Serina Proteases/genética , Complexo Shelterina , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Telomerase/química , Telomerase/genética , Telomerase/metabolismo , Telômero , Homeostase do Telômero , Proteínas de Ligação a Telômeros/química , Proteínas de Ligação a Telômeros/genéticaRESUMO
Numerous environmental carcinogens involve radical formation interacting with DNA to produce 2-deoxyribonolactone (dL), a major type of oxidized abasic site, implicated in DNA strand breaks, mutagenesis, and formation of covalent DNA-protein cross-links (DPC). Studies showed major dL-specific DPC occurred due to reactions with DNA polymerase beta (Polbeta) dependent on native conformation, while other DPC formed involved nonenzymatic reactions of DNA binding proteins with dL lesions. Polbeta appeared to play a major role in alleviating the cytotoxic effects of neocarzinostatin, which was used as a dL-producing agent. When a duplex DNA containing a dL at a site-specific position was incubated with purified histones, DPC were formed between dL and each histone protein, including H1, H2A, H2B, H3, and H4. Comparative kinetic analysis of DPC formation with histones and Polbeta revealed two distinct mechanisms of dL-mediated DPC formation. The rate of DPC formation with Polbeta was approximately two orders of magnitude higher than that with various histone proteins. These results indicate that catalytic activity of Polbeta mediates rapid DPC formation between dL and this DNA repair enzyme, whereas nonenzymatic reactions of dL with histones form DPC more slowly. The abundance of histones and their constant interaction with DNA may nevertheless yield significant levels of DPC with dL, as biomarkers of dL-induced cytotoxicity. Overall, data suggest that occurrence of dL-mediated DPC with histones may contribute to the genotoxic effects of dL in DNA.