Cloning and characterization of microRNAs from rainbow trout (Oncorhynchus mykiss): Their expression during early embryonic development

Raghuveer K Ramachandra,Scott Gahr,Caird E Rexroad,Mohamed Salem,Jianbo Yao

doi:10.1186/1471-213x-8-41

Abstract

BackgroundCurrent literature and our previous results on expression patterns of oocyte-specific genes and transcription factors suggest a global but highly regulated maternal mRNA degradation at the time of embryonic genome activation (EGA). MicroRNAs (miRNAs) are small, non-coding regulatory RNAs (19–23 nucleotides) that regulate gene expression by guiding target mRNA cleavage or translational inhibition. These regulatory RNAs are potentially involved in the degradation of maternally inherited mRNAs during early embryogenesis.ResultsTo identify miRNAs that might be important for early embryogenesis in rainbow trout, we constructed a miRNA library from a pool of unfertilized eggs and early stage embryos. Sequence analysis of random clones from the library identified 14 miRNAs, 4 of which are novel to rainbow trout. Real-time PCR was used to measure the expression of all cloned miRNAs during embryonic development. Four distinct expression patterns were observed and some miRNAs showed up-regulated expression during EGA. Analysis of tissue distribution of these miRNAs showed that some are present ubiquitously, while others are differentially expressed among different tissues. We also analyzed the expression patterns of Dicer, the enzyme required for the processing of miRNAs and Stat3, a transcription factor involved in activating the transcription of miR-21. Dicer is abundantly expressed during EGA and Stat3 is up-regulated before the onset of EGA.ConclusionThis study led to the discovery of 14 rainbow trout miRNAs. Our data support the notion that Dicer processes miRNAs and Stat3 induces expression of miR-21 and possibly other miRNAs during EGA. These miRNAs in turn guide maternal mRNAs for degradation, which is required for normal embryonic development.

Highlights

Current literature and our previous results on expression patterns of oocytespecific genes and transcription factors suggest a global but highly regulated maternal mRNA degradation at the time of embryonic genome activation (EGA)
To identify miRNAs that might be important for early embryogenesis in rainbow trout, we constructed a miRNA library from a pool of unfertilized eggs and early stage embryos
We present data to show that Dicer, the enzyme required for the processing of miRNAs, is abundantly expressed during EGA, and signal transducer and activator of transcription 3 (Stat3) which activates transcription of miRNA21, is up-regulated before the onset of EGA, supporting the important roles of these proteins in activation/ processing of miRNAs which in turn degrade maternal mRNAs during early embryonic development

Summary

Introduction

Current literature and our previous results on expression patterns of oocytespecific genes and transcription factors suggest a global but highly regulated maternal mRNA degradation at the time of embryonic genome activation (EGA). MicroRNAs (miRNAs) are small, non-coding regulatory RNAs (19–23 nucleotides) that regulate gene expression by guiding target mRNA cleavage or translational inhibition. The maternal-zygotic transition is characterized by asynchronous cell division, lengthening of cell cycles, cell motility and initiation of zygotic transcriptional machinery [1]. After this activation, the embryo becomes increasingly dependent on its own transcription and exhausts the maternally stored mRNAs. It was recently shown that it is critical to degrade maternally inherited mRNAs to achieve normal morphogenesis which is delayed if these inherited mRNAs are not degraded [4].

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Results

Conclusion