RESUMEN
BACKGROUND: The relapse rate in early stage non-small cell lung cancer (NSCLC) after surgical resection is high. Prognostic biomarkers may help identify patients who may benefit from additional therapy. The Helicase-like Transcription Factor (HLTF) is a tumor suppressor, altered in cancer either by gene hypermethylation or mRNA alternative splicing. This study assessed the expression and the clinical relevance of wild-type (WT) and variant forms of HLTF RNAs in NSCLC. METHODS: We analyzed online databases (TCGA, COSMIC) for HLTF alterations in NSCLC and assessed WT and spliced HLTF mRNAs expression by RT-ddPCR in 39 lung cancer cell lines and 171 patients with resected stage I-II NSCLC. RESULTS: In silico analyses identified HLTF gene alterations more frequently in lung squamous cell carcinoma than in adenocarcinoma. In cell lines and in patients, WT and I21R HLTF mRNAs were detected, but the latter at lower level. The subgroup of 25 patients presenting a combined low WT HLTF expression and a high I21R HLTF expression had a significantly worse disease-free survival than the other 146 patients in univariate (HR 1.96, CI 1.17-3.30; p = 0.011) and multivariate analyses (HR 1.98, CI 1.15-3.40; p = 0.014). CONCLUSION: A low WT HLTF expression with a high I21R HLTF expression is associated with a poor DFS.
Asunto(s)
Biomarcadores de Tumor/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Proteínas de Unión al ADN/genética , Recurrencia Local de Neoplasia/genética , Factores de Transcripción/genética , Adulto , Anciano , Empalme Alternativo/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Metilación de ADN/genética , Supervivencia sin Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Recurrencia Local de Neoplasia/patología , Estadificación de Neoplasias , PronósticoRESUMEN
MicroRNAs (miRNAs) are small, noncoding RNAs that negatively regulate gene expression. As miRNAs are involved in a wide range of biological processes and diseases, much effort has been invested in identifying their mRNA targets. Here, we present a novel combinatorial approach, RIP-chip-SRM (RNA-binding protein immunopurification + microarray + targeted protein quantification via selected reaction monitoring), to identify de novo high-confidence miRNA targets in the nematode Caenorhabditis elegans. We used differential RIP-chip analysis of miRNA-induced silencing complexes from wild-type and miRNA mutant animals, followed by quantitative targeted proteomics via selected reaction monitoring to identify and validate mRNA targets of the C. elegans bantam homolog miR-58. Comparison of total mRNA and protein abundance changes in mir-58 mutant and wild-type animals indicated that the direct bantam/miR-58 targets identified here are mainly regulated at the level of protein abundance, not mRNA stability.
Asunto(s)
Caenorhabditis elegans/genética , MicroARNs/metabolismo , Proteómica/métodos , Animales , Animales Modificados Genéticamente/genética , Animales Modificados Genéticamente/metabolismo , Caenorhabditis elegans/metabolismo , Cruzamientos Genéticos , Técnicas Inmunológicas/métodos , MicroARNs/genética , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Plásmidos/genética , Plásmidos/metabolismo , Biosíntesis de Proteínas , Interferencia de ARN , Estabilidad del ARN , ARN de Helminto/genética , ARN de Helminto/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , TransgenesRESUMEN
The soil nematode, Caenorhabditis elegans, occupies a central place in the short history of microRNA (miRNA) research. The converse is also true: miRNAs have emerged as key regulatory components in the life cycle of the worm, as well as numerous other organisms. Since the landmark discovery in 1993 of the first miRNA gene, lin-4, several other miRNAs have been characterized in detail in C. elegans and shown to participate in diverse biological processes. Moreover, the worm has provided, by virtue of its ease of genetic manipulation and amenability to high-throughput methods, an ideal platform for elucidating many general and conserved aspects of miRNA biology, namely mechanisms of biogenesis, target recognition, gene silencing, and regulation thereof. In this review, we summarize both the contribution of miRNAs to C. elegans physiology and development, as well as the contribution of C. elegans research to our understanding of general features of miRNA biology.