| A panel of candidate X-linked genes including BIRC4, BMP15 GAB3, HPRT1, JARID1C, MECP2, PLXNB3, RPS4X, SLC16A2, SLC25A6, UBE1, XIST and ZFX was selected to study X-linked gene expression and X inactivation in domestic cattle. Genes were selected by cDNA microarray analysis and a review of current scientific literature. Further analysis of transcript levels by quantitative PCR suggested escape from X inactivation in skeletal muscle for several genes. The same analysis in fibroblasts cultures showed changes in transcriptional activity, suggesting the influence of in vitro culture. Next, the transcriptional pattern of X-linked genes in pre-attachment bovine embryos derived in vivo and by two different in vitro production (IVP) systems was analyzed. There was an effect of the IVP system on transcription around the period of activation of the embryonic genome and marked differences between IVP and in vivo-derived embryos were also evident. We also investigated the transcriptional profile of X-linked genes in sexed pre-attachment embryos obtained by in vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT). Results showed dosage compensation between IVF-derived female and male blastocysts. Additionally, it was shown that X-linked gene expression was affected in SCNT-derived blastocysts, with female embryos showing severe disruption of their X-linked transcription. Finally, the transcriptional pattern of X-linked genes in a panel of organs from in vivo-derived fetuses and from SCNT-derived female conceptuses carrying an X/autosome translocation was determined. No significant differences were found in X-linked transcript levels for most organs between in vivo-derived female and male fetuses with the exception of HPRT1, RPS4X and SLC16A2 in skeletal muscle. The SCNT-derived females showed overall lower levels of transcripts than their in vivo-derived counterparts, with statistical significance for several transcripts various organs including muscle, placenta (cotyledon) and kidney. The results obtained in these studies suggest that X-linked gene expression in domestic cattle includes several genes escaping X-inactivation and that the transcriptional regulation of several of the genes included herein are subject to the influence of in vitro culture, probably through modulation of epigenetic modifications. |
