Laboratory of General Biology, Medical School, University of Ioannina , 45 110 Ioannina , Greece .
Stress. 2013 Nov;16(6):689-97. doi: 10.3109/10253890.2013.817554. Epub 2013 Jul 25.
Retrotransposons participate in cellular responses elicited by stress, and DNA methylation plays an important role in retrotransposon silencing and genomic imprinting during mammalian development. Assisted reproduction technologies (ARTs) may be associated with increased stress and risk of epigenetic changes in the conceptus. There are similarities in the nature and regulation of LTR retrotransposons and imprinted genes. Here, we investigated whether the methylation status of Human Endogenous Retroviruses (HERV)-K LTR retrotransposons and the imprinting signatures of the DLK1/MEG3. p57(KIP2) and IGF2/H19 gene loci are linked during early human embryogenesis by examining trophoblast samples from ART pregnancies and preimplantation genetic diagnosis (PGD) cases and matched naturally conceived controls. Methylation analysis revealed that HERV-Ks were totally methylated in the majority of controls while, in contrast, an altered pattern was detected in ART-PGD samples that were characterized by a hemi-methylated status. Importantly, DLK1/MEG3 demonstrated disturbed methylation in ART-PGD samples compared to controls and this was associated with altered HERV-K methylation. No differences were detected in p57(KIP2) and IGF2/H19 methylation patterns between ART-PGD and naturally conceived controls. Using bioinformatics, we found that while the genome surrounding the p57(KIP2) and IGF2/H19 genes differentially methylated regions had low coverage in transposable element (TE) sequences, the respective one of DLK1/MEG3 was characterized by an almost 2-fold higher coverage. Moreover, our analyses revealed the presence of KAP1-binding sites residing within retrotransposon sequences only in the DLK1/MEG3 locus. Our results demonstrate that altered HERV-K methylation in the ART-PGD conceptuses is correlated with abnormal imprinting of the DLK1/MEG3 locus and suggest that TEs may be affecting the establishment of genomic imprinting under stress conditions.
逆转录转座子参与细胞应激反应，DNA 甲基化在哺乳动物发育过程中的逆转录转座子沉默和基因组印迹中起着重要作用。辅助生殖技术（ART）可能与胚胎中应激增加和表观遗传变化的风险有关。LTR 逆转录转座子和印迹基因在性质和调控上有相似之处。在这里，我们通过检查来自 ART 妊娠和植入前遗传学诊断（PGD）病例以及匹配的自然受孕对照的滋养层样本，研究了人类内源性逆转录病毒（HERV）-K LTR 逆转录转座子的甲基化状态和印记特征是否与早期人类胚胎发生过程中的 DLK1/MEG3、p57(KIP2) 和 IGF2/H19 基因座相关。甲基化分析表明，在大多数对照中，HERV-K 完全甲基化，而在 ART-PGD 样本中检测到改变的模式，其特征是半甲基化状态。重要的是，与对照相比，ART-PGD 样本中的 DLK1/MEG3 表现出失调的甲基化，这与 HERV-K 甲基化改变有关。在 ART-PGD 和自然受孕对照之间，p57(KIP2) 和 IGF2/H19 甲基化模式没有差异。使用生物信息学，我们发现，虽然 p57(KIP2) 和 IGF2/H19 基因差异甲基化区域周围的基因组在转座元件（TE）序列中的覆盖度较低，但 DLK1/MEG3 的相应区域的覆盖度几乎高出两倍。此外，我们的分析还揭示了仅在 DLK1/MEG3 基因座中存在 KAP1 结合位点位于逆转录转座子序列内。我们的研究结果表明，ART-PGD 胚胎中 HERV-K 甲基化的改变与 DLK1/MEG3 基因座的异常印迹有关，并表明在应激条件下，TE 可能影响基因组印迹的建立。
