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Transcriptome high-throughput sequencing derived from mouse blastocysts with artificial oocyte activation treatment

ABSTRACT: Intracytoplasmic sperm injection (ICSI) has been an effective infertility treatment. Nevertheless, ICSI failures still occurred. One of the factors associated with ICSI failure is oocyte activation deficiency (AOD). The most commonly applied method of artificial oocyte activation (AOA) in humans includes ionophore. Although AOA is performed as a routine process in many assisted reproduction centers, up to date, there is no relevant study to expound whether AOA procedures increase developmental risks by disturbing subsequent gene expression during the embryonic development stages. Our results for the first time provide a profile of the changes in the global patterns of gene expression in AOA treatment versus ICSI generated mouse early embryos. In particular, we focus on the expression changes of imprinted genes. Another key observation in this study is that AOA treatment affects imprinted gene Igf2r expression and mehtylation, which is regulated by the imprinted Airn macro long non-coding (lnc) RNA.

ORGANISM(S): Mus musculus

PROVIDER: GSE155948 | GEO | 2021/11/10

REPOSITORIES: GEO

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Airn transcriptional overlap but not its lncRNA products induce imprinted Igf2r silencing.

Project description:To test if the imprinted long non-coding RNA (lncRNA) Airn transcription or its product silences the protein-coding Igf2r gene, we shortened the endogenous lncRNA to four different lengths by inserting polyadenylation (polyA) cassettes on the paternal chromosome via homologous recombination in ES cells. ES cell differentiation was used to recapitulate the developmental onset of Airn and Igf2r imprinted expression and polyA cassettes inserted before (T3, T16, T27) or after (T31, T51) the Igf2r promoter successfully truncated Airn, with the exception of T27. RNA hybridization to a tiling array (MIRTA) demonstrated loss of Airn upstream of the Igf2r promoter (except T27) and absence of novel spliced variants in all truncation alleles. To further test the transcriptional overlap model we moved the Airn promoter ~700bp before the Igf2r TSS in ES cells that lack an endogenous paternal Airn promoter and called this cell line FAP (Forward-Airn-Promoter). RNA hybridization to a tiling array (MIRTA) demonstrated that the moved Airn promoter expresses Airn in wildtype orientation and terminates at the wildtype 3' end. ES cell lines expressing different Airn variants were differentiated on gelatinized dishes after feeder cell depletion and LIF withdrawal by 0.27M-BM-5M retinoic acid for 5 days and Airn length was assayed.

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