Division of Veterinary Anatomy and Histology, Department of Animal Sciences, University of Milan, Milan, Italy.
Mol Hum Reprod. 2012 May;18(5):243-52. doi: 10.1093/molehr/gar077. Epub 2011 Dec 7.
Epigenetic modifications are established during gametogenesis and preimplantation embryonic development. Any disturbance of the normal natural environment during these critical phases could cause alterations of the epigenetic signature. Histone acetylation is an important epigenetic modification involved in the regulation of chromatin organization and gene expression. The present study was aimed to determine whether the proper establishment of post-translational histone H4 acetylation at lysine 8 (AcH4K8), 12 (AcH4K12) and 16 (AcH4K16) of equine oocytes is adversely affected during in vitro maturation (IVM) when compared with in vivo matured oocytes collected from naturally cycling mares not undergoing ovarian hyperstimulation. The acetylation patterns were investigated by means of indirect immunofluorescence staining with specific antibodies directed against the acetylated lysine residues. Our results indicate that the acetylation state of H4 is dependent on the chromatin configuration in immature germinal vesicle (GV) stage oocytes and it changes in a residue-specific manner along with the increase of chromatin condensation. In particular, the levels of AcH4K8 and AcH4K12 increased significantly, while AcH4K16 decreased significantly from the fibrillar to the condensed state of chromatin configuration within the GV. Moreover, during meiosis, K8 and K12 were substantially deacetylated without any differences between in vivo and in vitro conditions, while K16 displayed a strong acetylation in oocytes matured in vivo, and in contrast, it was markedly deacetylated following IVM. Although the functional meaning of residue-specific acetylation during oocyte differentiation and meiotic resumption needs further investigation, our results support the hypothesis that IVM conditions can adversely affect oocyte ability to regulate the epigenetic reprogramming, critical for successful meiosis and subsequent embryonic development.
表观遗传修饰是在配子发生和胚胎植入前发育过程中建立的。在这些关键阶段，正常自然环境的任何干扰都可能导致表观遗传特征的改变。组蛋白乙酰化是一种重要的表观遗传修饰，参与调节染色质结构和基因表达。本研究旨在确定体外成熟（IVM）过程中，是否会对马卵母细胞中转录后组蛋白 H4 赖氨酸 8（AcH4K8）、12（AcH4K12）和 16（AcH4K16）的乙酰化修饰产生不利影响，与未接受卵巢过度刺激的自然周期母马体内成熟的卵母细胞相比。通过针对乙酰化赖氨酸残基的特异性抗体的间接免疫荧光染色来研究乙酰化模式。我们的结果表明，H4 的乙酰化状态依赖于不成熟生发泡（GV）阶段卵母细胞的染色质构型，并且随着染色质浓缩程度的增加，以残基特异性的方式发生变化。特别是，AcH4K8 和 AcH4K12 的水平显著增加，而 AcH4K16 的水平则从纤维状到浓缩的染色质构型显著降低。此外，在减数分裂过程中，K8 和 K12 被大量去乙酰化，体内和体外条件之间没有差异，而 K16 在体内成熟的卵母细胞中表现出强烈的乙酰化，相反，在 IVM 后明显去乙酰化。虽然卵母细胞分化和减数恢复过程中特定残基乙酰化的功能意义需要进一步研究，但我们的结果支持这样一种假设，即 IVM 条件可能会对卵母细胞调节表观遗传重编程的能力产生不利影响，这对于成功的减数分裂和随后的胚胎发育至关重要。
