MicroRNA Expression during Bovine Oocyte Maturation and Fertilization.

Graham C Gilchrist,Daniele Merico,Allison Tscherner,Jonathan Lamarre,Thomas Nalpathamkalam

doi:10.3390/ijms17030396

Graham C Gilchrist, Daniele Merico + Show 3 more

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https://doi.org/10.3390/ijms17030396

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Abstract

Successful fertilization and subsequent embryo development rely on complex molecular processes starting with the development of oocyte competence through maturation. MicroRNAs (miRNAs) are small non-coding RNA molecules that function as gene regulators in many biological systems, including the oocyte and embryo. In order to further explore the roles of miRNAs in oocyte maturation, we employed small RNA sequencing as a screening tool to identify and characterize miRNA populations present in pools of bovine germinal vesicle (GV) oocytes, metaphase II (MII) oocytes, and presumptive zygotes (PZ). Each stage contained a defined miRNA population, some of which showed stable expression while others showed progressive changes between stages that were subsequently confirmed by quantitative reverse transcription polymerase chain reaction (RT-PCR). Bta-miR-155, bta-miR-222, bta-miR-21, bta-let-7d, bta-let-7i, and bta-miR-190a were among the statistically significant differentially expressed miRNAs (p < 0.05). To determine whether changes in specific primary miRNA (pri-miRNA) transcripts were responsible for the observed miRNA changes, we evaluated pri-miR-155, -222 and let-7d expression. Pri-miR-155 and -222 were not detected in GV oocytes but pri-miR-155 was present in MII oocytes, indicating transcription during maturation. In contrast, levels of pri-let-7d decreased during maturation, suggesting that the observed increase in let-7d expression was likely due to processing of the primary transcript. This study demonstrates that both dynamic and stable populations of miRNAs are present in bovine oocytes and zygotes and extend previous studies supporting the importance of the small RNA landscape in the maturing bovine oocyte and early embryo.

Highlights

Dynamic changes in the mRNAs and proteins expressed in oocytes prior to fertilization represent an important component of successful mammalian embryo development
In order to investigate the potential biological roles of miRNAs identified in our datasets, we cross-referenced predicted mRNA targets derived from TargetScan with the data from the oocyte and zygote proteome signature recently published by Deutsch et al [30]
Among the proteins that were detected in the proteome, the expression changes from metaphase II (MII) to zygotes corresponded with high frequency to the changes expected based on our miRNA results, namely, that increasing expression of any given miRNA will decrease the expression of the protein encoded by its target and that decreasing miRNA expression will have the opposite effect

Summary

Introduction

Dynamic changes in the mRNAs and proteins expressed in oocytes prior to fertilization represent an important component of successful mammalian embryo development. Pre-ovulatory germinal vesicle (GV) oocytes remain arrested in prophase I of meiosis until a surge of gonadotropins triggers nuclear and cytoplasmic maturation events [1]. The oocyte resumes meiosis by undergoing germinal vesicle breakdown (GVBD), extrusion of the first polar body, chromatin remodelling and cytoplasmic organelle reorganization before it is arrested again in metaphase II (MII), the point at which it becomes competent for fertilization [1,2]. Oocyte maturation is directed in part by specific patterns of gene expression [3], even though it is generally accepted that transcription in oocytes is markedly decreased beyond the GVBD stage. Temporal changes in the transcripts present during oocyte maturation have been recognized [7], and such changes may be due to microRNA (miRNA)-mediated regulation of transcript levels [8,9]

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