小鼠早期胚胎发育过程中的DNA去甲基化

表观遗传修饰在基因转录与表达、细胞生长与分化以及动物个体正常发育等过程中都具有重要的调控作用。表观遗传修饰发生异常,会引起机体生长发育中的各种异常。哺乳动物从精卵受精到附植前的胚胎早期发育阶段会发生重要的表观遗传重编程,主要包括DNA甲基化和组蛋白修饰。精卵受精后DNA发生主动和被动2种方式的去甲基化。本文主要综述了与DNA甲基化相关的蛋白和早期胚胎发育过程中的去甲基化机制,并对小鼠附植前胚胎发育过程中的DNA甲基化的动态变化进行了详细的论述。     

早期胚胎DNA甲基化研究进展

DNA甲基化修饰是研究最多的表观遗传修饰之一,在调控基因转录、染色体结构稳定、基因印迹、X染色体失活等方面发挥作用。尽管DNA甲基化是一种稳定的修饰,但其在个体发育进程中是动态变化的。目前,人们对早期胚胎发育中DNA甲基化修饰研究还不全面,随着全基因组DNA甲基化分析技术的进步,其在早期胚胎中的功能也逐渐揭示。作者主要论述了DNA甲基转移酶(DNMTs)的发现及其调控作用和DNA甲基化在早期胚胎中的作用。     

早期胚胎DNA甲基化研究进展

DNA甲基化修饰是研究最多的表观遗传修饰之一，在调控基因转录、染色体结构稳定、基因印迹、X染色体失活等方面发挥作用。尽管DNA甲基化是一种稳定的修饰，但其在个体发育进程中是动态变化的。目前，人们对早期胚胎发育中DNA甲基化修饰研究还不全面，随着全基因组DNA甲基化分析技术的进步，其在早期胚胎中的功能也逐渐揭示。作者主要论述了DNA甲基转移酶（DNMTs）的发现及其调控作用和DNA甲基化在早期胚胎中的作用。     

DNA甲基化/去甲基化与哺乳动物胚胎的体外发育

DNA甲基化及去甲基化是哺乳动物表观遗传修饰的主要方式之一,与哺乳动物胚胎的发育密切相关。因此深入研究DNA甲基化与去甲基化的发生机制,对于改善早期胚胎的发育具有重要意义。本文对哺乳动物胚胎早期发育过程中的DNA甲基化动态修饰进行了综述。     

DNA甲基化在哺乳动物卵母细胞和早期胚胎发育中的研究进展

DNA甲基化(DNA methylation)是一种动态、可逆并可以遗传的表观遗传修饰模式,主要发生在哺乳动物原始生殖细胞和早期胚胎发育过程中,能够通过高动态和协同的核酶网络附着在DNA的CpG区域,同时还通过改变调控区域的功能状态进而调控基因表达且不影响DNA序列所携带的遗传信息。DNA甲基化主要涉及基因组印迹、转座元件沉默、X染色体失活和衰老等多种关键生理过程,在哺乳动物卵母细胞和胚胎发育中发挥着重要作用。本文介绍了DNA甲基化的建立与去除机制及其生物学功能,重点阐述了DNA甲基化在哺乳动物卵母细胞和胚胎发育过程中精准生成、维持、读取和删除等动态变化过程,为进一步研究哺乳动物表观遗传调控提供参考依据。     

DNA甲基化动态模式在重编码过程中的作用

DNA甲基化作为主要的表观遗传修饰方式在细胞分裂与分化过程中扮演着重要角色。在生殖细胞生成与胚胎的早期发育过程中,DNA甲基化具有动态变化的特征,其动态模式直接影响细胞核重编码程序的,控制胚胎的成活率及发育状况。本文综述了DNA甲基化修饰在核重编程过程中的动态模式、相关酶及甲基化结合蛋白的作用模式等。     

组蛋白H3K4me3甲基化修饰与哺乳动物早期胚胎发育

表观遗传调控是细胞分化过程中的主要机制之一,尤其在生殖细胞分化调控中尤为重要。而组蛋白甲基化修饰是表观遗传信息的重要载体和生命活动的重要调控因子,对细胞的状态和胚胎的发生与发育具有决定性的作用,就组蛋白H3K4me3甲基化修饰与哺乳动物早期胚胎发育研究进展进行了综述。     

DNA甲基化在猪胚胎发育过程中的研究进展

猪的胚胎发育需要经历受精、卵裂、孵化、形态转变、附植、器官分化等一系列重要的生理阶段。虽然在胚胎发育过程中基因的严格表达与正确指导是胚胎能否正常发育的决定性条件,但研究表明DNA甲基化修饰对胚胎的发育也起着必不可少的作用。DNA甲基化是一种常见且重要的表观遗传修饰,虽然不改变DNA的一级序列,但也包含可遗传信息,并在基因的转录调控中起重要作用。在猪的胚胎发育中,DNA甲基化呈现出高度动态的过程,这一过程受孕期母体营养和发育环境条件影响。本文将从胚胎早期发育、体细胞核移植和孕期母体营养三个方面来阐述DNA甲基化对胚胎发育的影响,为进一步研究猪胚胎在发育过程中的DNA甲基化机制和提高体细胞核移植的成功率提供参考。     

DNA甲基化的表观遗传机制及其在动物遗传育种中应用

表观遗传是一类没有产生DNA碱基序列的改变但表型特征却发生变化的调控机制,同时这种变化是可以遗传给后代的。DNA甲基化是DNA分子上的胞嘧啶在DNA甲基转移酶的作用下与一个甲基基团共价结合,被修饰为5-甲基胞嘧啶,进而调控基因表达的一种表观遗传形式。多项研究表明,DNA甲基化对动物的抗病性能、生产性能、繁殖性能均有不同程度的影响,因此探明DNA甲基化的表观遗传机制,能够为分子育种提供理论依据,将有助于更好更快的选育新品种或品系,并充分发挥其遗传潜力。     

表观遗传修饰对转基因和克隆胚早期发育的影响研究进展

表观遗传修饰在基因表达和克隆胚的早期发育方面有重要作用.本文从DNA甲基化、组蛋白修饰、染色质重塑和非编码RNA调控方面综述了表观遗传的发生机制及其对转基因和克隆胚早期发育的影响.该文对表观遗传的研究有重要的指导作用.     

DNA methylation status of bovine blastocyst embryos obtained from various procedures

DNA methylation is an important factor for the regulation of gene expression in early embryos. It is well known that the satellite I sequence is more heavily methylated in bovine somatic cell nuclear transfer (NT-SC) embryos than in embryos derived from in vitro fertilization (IVF). However, the methylation status of bovine embryos obtained by other procedures is not well known. To clarify DNA methylation levels of bovine embryos obtained from various procedures, we examined satellite I sequences in bovine blastocyst (BC) embryos derived from NT-SC, NT using embryonic blastomeres (NT-EM), in vivo (Vivo), IVF and parthenogenetic treatment (PA). Furthermore, in order to evaluate the efficacy of DNA demethylation by the NT procedure, we determined the DNA methylation levels in bovine embryos in which NT was recapitulated (Re-NT). Although the DNA methylation levels in the NT-SC embryos were higher than those in the other embryos, the NT-EM embryos exhibited lower DNA methylation levels. The satellite I sequence in the NT-SC embryos was more demethylated than that in the donor cells. Although the DNA methylation level in the individual NT-SC embryos showed variation, the full-term developmental efficacy of these embryos were not different. These findings suggest that the methylation level of the satellite I sequence at the BC stage is not related to the abnormalities of bovine embryos produced by NT-SC. There was no difference in methylation levels between Re-NT and NT-SC embryos. Our results indicated that the DNA methylation status differed among embryos produced by various methods and that at least some of the demethylation of the donor cell genome occurred in the recipient cytoplast after NT-SC, but the demethylation ability of the NT procedure was noted in the first NT but not in the second NT.     

Expression Analysis of Autophagy Regulators Atg5 and Beclin1 during Early Embryonic Development of Mouse (Mus musculus) from Different Sources

This study aimed to explore the expression patterns of autophagy regulators Atg5 and Beclin1 in the early embryonic development and the effects of different embryonic production methods on the expression of the two factors. Female mice aged 6-8 weeks were subjected to superovulation and divided into 2 groups. The mouse oocytes of one group were collected, and cultured in vitro after parthenogenetic activation. The other group of female mice were caged with male mice (1:1), and the next day, the mouse fertilized eggs were collected for in vitro culture. Parthenogenetic activated embryos and naturally fertilized embryos were collected at 2 cell stage, 4-8 cell stage, mulberry embryo stage and blastocyst stage, respectively. RNA and protein were extracted, real-time fluorescence quantitative PCR, Western blot and other methods were used to detect the expression of key autophagy factors Atg5 and Beclin1. And indirect immunofluorescence was used to detect the expression and location of Atg5 and Beclin1 in mouse blastocysts. The results showed that Atg5 and Beclin1 were expressed in all development stages of naturally fertilized and parthenogenetic activated embryos in mice, and showed a high level in the early stage of embryonic development. The expression of Atg5 and Beclin1 were gradually reduced from the 2 cell stage in mouse naturally fertilized embryos. The expression levels of Atg5 and Beclin1 in parthenogenetic activated embryos were the highest in the 4-8 cell stage, which was extremely significantly different from the naturally fertilized embryos of the same period (P<0.01). From the 4 cell stage, the expression levels of Atg5 and Beclin1 in parthenogenetic activated embryos were higher than naturally fertilized embryos at all subsequent stages, the difference was extremely significantly different (P<0.01). In mouse blastocysts, the fluorescence of Atg5 and Beclin1 protein could be detected in the trophoblast cells and the inner cell mass, but the fluorescence intensity in the inner cell mass was higher than that in the trophoblast cells. In addition, the fluorescence intensity of Beclin1 protein in the inner cell mass of parthenogenetic activated embryos was higher than that in naturally fertilized embryos. Atg5 and Beclin1, the key autophagy factors, are expressed at different levels in the early development of mouse embryos from different sources. It is suggested that the regulation of autophagy on early embryonic development is related to embryo production modes. The results will provide a theoretical basis for further exploring the role of autophagy in the physiological regulation of mammalian embryo development.     

自噬调节因子Atg5和Beclin1在不同来源小鼠胚胎早期发育过程中的表达分析

旨在探究自噬调节因子Atg5和Beclin1在胚胎早期发育过程中的表达模式及胚胎的不同生产方式对两种因子表达的影响。本研究将6~8周龄雌性小鼠进行超数排卵,分为2组,一组收集小鼠卵母细胞,孤雌激活处理后进行体外培养;另一组超排小鼠与公鼠1：1合笼,第2天收集小鼠受精卵进行体外培养;分别在2细胞期、4~8细胞期、桑葚胚期和囊胚期收集不同阶段小鼠孤雌激活胚胎和自然受精胚胎。提取RNA和蛋白,通过实时荧光定量PCR、Western blot等方法检测自噬关键因子Atg5和Beclin1的表达,通过间接免疫荧光法检测Atg5和Beclin1在小鼠囊胚中的表达定位。结果显示,小鼠自然受精和孤雌激活胚胎在发育各时期均可表达Atg5和Beclin1,表达量在胚胎发育的早期呈现出较高的水平,其中二者的表达在小鼠自然受精胚胎中从2细胞期起逐渐降低,而在孤雌激活胚胎的4~8细胞阶段表达量最高,与同期自然受精胚胎差异极显著（P<0.01）;从4细胞期开始,各时期孤雌激活胚胎中Atg5和Beclin1蛋白表达水平均高于自然受精胚胎,差异极显著（P<0.01）;在囊胚中,滋养层细胞和内细胞团中均可检测到Atg5和Beclin1蛋白的荧光,但内细胞团中的荧光强度高于滋养层细胞,且Beclin1蛋白在孤雌激活胚胎囊胚内细胞团中荧光强度高于自然受精胚胎。自噬关键因子Atg5和Beclin1在不同来源小鼠胚胎早期发育各时期均有不同程度的表达,提示自噬对早期胚胎发育的调控作用与胚胎的生产方式存在一定关联,研究结果为进一步探索细胞自噬参与哺乳动物胚胎发育的生理调控提供理论依据。     

Epigenetic characteristics of cloned and in vitro-fertilized swamp buffalo (Bubalus bubalis) embryos

Swamp buffalos are becoming endangered due to reproductive inefficiencies. This is of concern because many countries depend heavily on their products. Somatic cell nuclear transfer (SCNT) is a potential strategy for preserving endangered species. To date, SCNT in swamp buffalo has succeeded in the creation of blastocyst embryos. However, development to term of SCNT swamp buffalos is extremely limited, and only 1 live birth has been reported. An abnormal epigenetic mechanism is suspected to be the cause of developmental failure, as is also seen in other species. The DNA methylation and histone acetylation are key players in epigenetic modification and display marked variability during embryonic preimplantation development. Knowledge of epigenetic modifications will aid in solving the developmental problems of SCNT embryos and improving reproductive technology in the swamp buffalo. The objective of this study was to determine the relationship between preimplantation embryonic development and 2 epigenetic patterns, global DNA methylation and histone acetylation, in SCNT and in vitro-fertilized (IVF) swamp buffalo embryos. In addition, we examined the correlations between those 2 mechanisms in the SCNT and IVF swamp buffalo embryos throughout the developmental stages using double immunostaining and quantification of the emission intensities using confocal microscopy. We discovered an aberrant methylation pattern in early preimplantation-stage swamp buffalo SCNT embryos. In addition, greater variability in the DNA methylation levels among nuclei within SCNT embryos was discovered. Hyperacetylation was also observed in SCNT embryos compared with IVF embryos at the 4- and 8-cell stages (P < 0.05). Dynamic changes and interplay between these 2 epigenetic mechanisms could be crucial for embryonic development during the early preimplantation period. The aberrancies uncovered here may contribute to the low efficiency of SCNT.     

胚胎母源-合子转换期中的表观遗传调控

哺乳动物个体由终端分化的单倍体精子和卵子受精融合成的双倍体受精卵发育而来。在胚胎发育的初始阶段,合子基因组处于休眠状态,胚胎发育调控由卵母细胞内母源调控逐渐转换为合子基因组调控(Maternal-Zygotic Transition,MZT)。在此期间,随着母源物质的清除,合子基因组激活(Zygotic Genome Activation,ZGA),但调控MZT的具体分子机制还不清楚。最新研究表明,DNA甲基化、染色质重塑、组蛋白表观修饰、ncRNA在MZT和ZGA中发挥重要的作用。本文总结了上述4种表观修饰在动物植入前胚胎MZT中的生物学功能和特异的分子机制,对揭示动物胚胎MZT的调控机理有借鉴意义。