RESUMEN
MicroRNA expression is important for gene regulation and deregulated microRNA expression is often observed in diseases such as cancer. The processing of primary microRNA transcripts is an important regulatory step in microRNA biogenesis. Due to low expression level and association with chromatin, primary microRNAs are challenging to study in clinical samples where input material is limited. Here, we present a high-sensitivity targeted method to determine processing efficiency of several hundred primary microRNAs from total RNA that requires relatively few RNA sequencing reads. We validate the method using RNA from HeLa cells and show the applicability to clinical samples by analyzing RNA from normal liver and hepatocellular carcinoma. We identify 24 primary microRNAs with significant changes in processing efficiency from normal liver to hepatocellular carcinoma, among those the highly expressed miRNA-122 and miRNA-21, demonstrating that differential processing of primary microRNAs is occurring and could be involved in disease. With our method presented here we provide means to study pri-miRNA processing in disease from clinical samples.
Asunto(s)
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , MicroARNs/genética , Análisis de Secuencia de ARN/métodos , Regulación Neoplásica de la Expresión Génica , Células HeLa , Secuenciación de Nucleótidos de Alto Rendimiento , HumanosRESUMEN
MAIN CONCLUSION: Our work provides a survey of mature miRNAs, their target genes and primary precursors identified by in-silico approach in leaf transcriptomes of five selected Hypericum species. MiRNAs are small non-coding RNA molecules found in animals, terrestrial plants, several algae and molds. As their role lies in the post-transcriptional gene silencing, these tiny molecules regulate many biological processes. Phyto-miRNAs are considered the important regulators of secondary metabolism in medicinal plants. The genus Hypericum comprises many producers of bioactive compounds, mainly unique naphtodianthrones with a great therapeutic potential. The main goal of our work was to identify genetically conserved miRNAs, characterize their primary precursors and target sequences in the leaf transcriptomes of five Hypericum species using in-silico approach. We found 20 sequences of potential Hypericum pri-miRNAs, and predicted and computationally validated their secondary structures. The mature miRNAs were identified by target genes screening analysis. Whereas predicted miRNA profiles differed in less genetically conserved families, the highly conserved miRNAs were found in almost all studied species. Moreover, we detected several novel highly likely miRNA-mRNA interactions, such as mir1171 with predicted regulatory role in the biosynthesis of melatonin in plants. Our work contributes to the knowledge of Hypericum miRNAome and miRNA-mRNA interactions.
Asunto(s)
Biología Computacional , Hypericum/genética , MicroARNs/genética , Hojas de la Planta/genética , Transcriptoma/genética , Regulación de la Expresión Génica de las Plantas , MicroARNs/química , MicroARNs/metabolismo , Conformación de Ácido Nucleico , Proyectos PilotoRESUMEN
microRNAs (miRNAs or miRs) are small non-coding RNAs that are involved in post-transcriptional regulation of their target genes in a sequence-specific manner. Emerging evidence demonstrates that miRNAs are critical regulators of lipid synthesis, fatty acid oxidation and lipoprotein formation and secretion. Dysregulation of miRNAs disrupts gene regulatory network, leading to metabolic syndrome and its related diseases. In this review, we introduced epigenetic and transcriptional regulation of miRNAs expression. We emphasized on several representative miRNAs that are functionally involved into lipid metabolism, including miR-33/33(â), miR122, miR27a/b, miR378/378(â), miR-34a and miR-21. Understanding the function of miRNAs in lipid homeostasis may provide potential therapeutic strategies for fatty liver disease.