The combination of transcriptomics and informatics identifies pathways targeted by miR-204 during neurogenesis and axon guidance.

Ivan Conte,Jose Manuel Garcia-Manteiga,Dejan Lazarevic,Sandro Banfi,Chiara Migliore,Germana Meroni,Nathan Paul Davidson,Sabrina Carrella,Remo Sanges,Paola Bovolenta,Raffaella Avellino,Nicholas Bockett,Warren Emmett,Raquel Marco-Ferreres,Elia Stupka,Stefania Merella,David Van Heel

doi:10.1093/nar/gku498

Abstract

Vertebrate organogenesis is critically sensitive to gene dosage and even subtle variations in the expression levels of key genes may result in a variety of tissue anomalies. MicroRNAs (miRNAs) are fundamental regulators of gene expression and their role in vertebrate tissue patterning is just beginning to be elucidated. To gain further insight into this issue, we analysed the transcriptomic consequences of manipulating the expression of miR-204 in the Medaka fish model system. We used RNA-Seq and an innovative bioinformatics approach, which combines conventional differential expression analysis with the behavior expected by miR-204 targets after its overexpression and knockdown. With this approach combined with a correlative analysis of the putative targets, we identified a wider set of miR-204 target genes belonging to different pathways. Together, these approaches confirmed that miR-204 has a key role in eye development and further highlighted its putative function in neural differentiation processes, including axon guidance as supported by in vivo functional studies. Together, our results demonstrate the advantage of integrating next-generation sequencing and bioinformatics approaches to investigate miRNA biology and provide new important information on the role of miRNAs in the control of axon guidance and more broadly in nervous system development.

Highlights

MicroRNAs, a class of short non-coding RNAs (∼22 nucleotides in length), have emerged as important regulators of gene expression in development [1]
Our results demonstrate the advantage of integrating next-generation sequencing and bioinformatics approaches to investigate miRNA biology and provide new important information on the role of miRNAs in the control of axon guidance and more broadly in nervous system development
They exert their function in animal cells by binding, with imperfect base pairing, to target sites in the 3 un-translated regions (3 UTRs) of messenger RNAs

Summary

Introduction

MicroRNAs (miRNAs), a class of short non-coding RNAs (∼22 nucleotides in length), have emerged as important regulators of gene expression in development [1]. Each miRNA is estimated to regulate, on average, the expression of 100–200 distinct target genes, so that the whole miRNA apparatus seems to participate in the control of gene expression for a significant proportion of the vertebrate transcriptome [6,7]. MiRNAs are as important as transcription factors or signaling molecules in controlling cellular processes. Recent discoveries have revealed a model in which miRNA regulatory events are woven into the known transcription factor and signaling networks that control cell fate and differentiation, modulating their activity through positive and negative feedback loops to promote programs that define the fate and character of

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Conclusion