Abstract
BackgroundThe expression of microRNAs (miRNAs) is essential for the proper development of the mammalian embryo. A maternal exposure to endocrine disrupting chemicals during preimplantation bears the potential for transgenerational inheritance of disease through the epigenetic perturbation of the developing embryo. A comprehensive assembly of embryo-specific miRNAs and respective isoforms (isomiR) is lacking to date. We aimed at revealing the sex-specific miRNA expression profile of single porcine blastocysts developing in gilts orally exposed to exogenous estradiol-17 β (E2). Therefore we analyzed the miRNA profile specifically focusing on isomiRs and potentially embryo-specific miRNAs.ResultsDeep sequencing of small RNA (small RNA-seq) result in the detection of miRNA sequences mapping to known and predicted porcine miRNAs as well as novel miRNAs highly conserved in human and cattle. A set of highly abundant miRNAs and a large number of rarely expressed miRNAs were identified by using a small RNA analysis pipeline, which was integrated into a novel Galaxy workflow specifically benefits incompletely annotated species. In particular, orthologue species information increased the total number of annotated miRNAs, while mapping to other non-coding RNAs avoided falsely annotated miRNAs. Neither the low nor the high dose of E2 treatment (10 and 1000 µE2/kg body weight daily, respectively) affected the miRNA profile in blastocysts despite the distinct differential mRNA expression and DNA methylation found in previous studies. The high number of generated sequence reads enabled a comprehensive analysis of the isomiR repertoire showing various templated and non-templated modifications. Furthermore, potentially blastocyst-specific miRNAs were identified.ConclusionsIn pre-implantation embryos, numerous distinct isomiRs were discovered indicating a high complexity of miRNA expression. Neither the sex of the embryo nor a maternal E2 exposure affected the miRNA expression profile of developing porcine blastocysts. The adaptation to the continuous duration of the E2 treatment might explain the lack of an effect.
Highlights
The expression of microRNAs is essential for the proper development of the mammalian embryo
Endocrine disrupting chemicals (EDCs) are natural and synthetic substances that may impact on health by deteriorating the endogenous hormone system [1, 2]
Estradiol-17β (E2) is the most potent endogenous estrogen in the body. Because it is present in the environment and can adversely act on the developing organisms, it is considered as an EDC [6, 7]
Summary
The expression of microRNAs (miRNAs) is essential for the proper development of the mammalian embryo. We aimed at revealing the sex-specific miRNA expression profile of single porcine blastocysts developing in gilts orally exposed to exogenous estradiol-17β (E2). Orthologue species information increased the total number of annotated miRNAs, while mapping to other non-coding RNAs avoided falsely annotated miRNAs. Neither the low nor the high dose of E2 treatment (10 and 1000 μ E2/kg body weight daily, respectively) affected the miRNA profile in blastocysts despite the distinct differential mRNA expression and DNA methylation found in previous studies. Neither the sex of the embryo nor a maternal E2 exposure affected the miRNA expression profile of developing porcine blastocysts. Estradiol-17β (E2) is the most potent endogenous estrogen in the body Because it is present in the environment and can adversely act on the developing organisms, it is considered as an EDC [6, 7]
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