Environmental influence on gametes and embryos

哺乳动物受精的一个普遍特征是，受精的精子会在卵子中引发一系列重复的钙振荡，这些振荡持续几个小时，最终形成原核。小鼠的这种钙振荡模式对于受精后卵子激活的早期事件和足月宫内发育都是必不可少的。精子内导致这些钙振荡的因素是一种睾丸特异性磷脂酶C，PLC Zeta，它在精子-卵子质膜融合后从精子头部释放出来，迄今在所有研究的哺乳动物精子中都发现了这种磷脂酶C，包括人类。钙的振荡也受鸡蛋内部因素的控制。正在使用小鼠模型进行研究，以检查卵子内负责控制钙振荡行为和钙再吸收的分子，这些过程对于受精时钙振荡的持续至关重要。我们发现，在小鼠卵母细胞成熟或受精过程中，体细胞钙进入的主要机制，即所谓的储备式钙进入，并不是钙进入所必需的。取而代之的是使用替代通道，包括电压操作的钙通道CaV3.2和瞬时受体电位通道TRPM7。我们还发现，来自不同普通实验室小鼠品系的卵子在受精后第一次钙瞬变的持续时间和最大幅度、振荡频率和钙储存方面存在显著差异。这些钙振荡模式的差异是由于不同品系小鼠的卵细胞因子不同造成的。我们的结果支持了鸡蛋的固有特性在决定钙振荡模式中的重要性，并对解释和比较受精时钙动力学的研究具有重要意义。在我们正在进行的研究中，我们正在确定受精后钙进入卵子的其他机制。我们预计，通过更好地了解卵子激活过程中钙振荡行为的分子和细胞调控模式，我们可以了解环境因素和疾病状态是如何改变早期胚胎发育的。 许多必需的分子是由母体mRNAs编码的，这些分子在卵母细胞成熟之前处于休眠状态，但对受精和胚胎发育的早期阶段至关重要。其中一个步骤，胚胎基因组激活(EGA)，是由卵母细胞成熟过程中启动的内在发育程序协调的。我们发表的工作表明，tankyrase是一种调节β-catenin水平的多聚(ADP-核糖基)聚合酶，在卵母细胞成熟过程中经历程序性翻译，在小鼠EGA中发挥重要作用。新翻译的tankyrase触发了Axin的蛋白酶体降解，减少了对β-catenin的定向破坏，并促进了β-catenin介导的靶基因的转录，包括Myc。MYC在2-细胞胚胎中介导核糖体RNA转录，支持全球蛋白质合成。使用基因敲除或化学抑制抑制tankyrase活性会导致核β-连环蛋白的丢失，并导致转录和组蛋白H3乙酰化的整体减少。染色质和转录图谱表明，发育停滞在2-细胞中期之前，部分是由β-连环蛋白和MYC的减少所调节的。这些发现表明，Tankyrase的转录后调控是翻译后连环蛋白激活的一种不依赖配体的发育机制，是完成EGA所必需的。这些研究揭示了暴露在环境化学品中可能会扰乱并可能影响人类生育能力的基本遗传过程。

A universal feature of fertilization in mammals is that the fertilizing sperm evokes a series of repetitive calcium oscillations in the egg that persist for several hours and terminate with pronucleus formation. This pattern of calcium oscillations in mice is essential for both early events of egg activation in response to fertilization and for full term intrauterine development to occur. The factor within sperm responsible for inducing these calcium oscillations is a testis-specific phospholipase C, PLC zeta, which is released from the sperm head after sperm-egg plasma membrane fusion and is found in all mammalian sperm studied to date, including human. Calcium oscillations are also controlled by factors within the egg. Studies are being performed using the mouse model to examine molecules within the egg that are responsible for controlling calcium oscillation behavior and calcium reuptake, processes that are essential for the continuation of calcium oscillations at fertilization. We found that the major mechanism of calcium entry in somatic cells, known as store-operated calcium entry, is not necessary for calcium entry during oocyte maturation or at fertilization in mice. Instead, alternate channels are utilized including the voltage operated calcium channel, CaV3.2, and the transient receptor potential channel, TRPM7. We also found that eggs from different common laboratory mouse strains have significant differences in the duration and maximum amplitude of the first calcium transient following fertilization, frequency of oscillations, and calcium stores. These differences in calcium oscillation patterns result from variation in egg factors across different mouse strains. Our results support the importance of egg-intrinsic properties in determining calcium oscillation patterns and have important implications for the interpretation and comparison of studies on calcium dynamics at fertilization. In our ongoing studies, we are determining additional mechanisms by which calcium enters the egg following fertilization. We anticipate that by achieving a better understanding of the molecular and cellular modes of regulation of calcium oscillatory behavior during egg activation, we can learn how early embryo development is altered by environmental factors and by disease states. A number of essential molecules are encoded by maternal mRNAs that are dormant until oocyte maturation but critically important for fertilization and the early steps of embryonic development. One of these steps, embryonic genome activation (EGA), is orchestrated by an intrinsic developmental program initiated during oocyte maturation. We published work showing that tankyrase, a poly(ADP-ribosyl) polymerase that regulates beta-catenin levels, undergoes programmed translation during oocyte maturation and serves an essential role in mouse EGA. Newly translated tankyrase triggers proteasomal degradation of axin, reducing targeted destruction of beta-catenin and promoting beta-catenin-mediated transcription of target genes, including Myc. MYC mediates ribosomal RNA transcription in 2-cell embryos, supporting global protein synthesis. Suppression of tankyrase activity using knockdown or chemical inhibition causes loss of nuclear beta-catenin and global reductions in transcription and histone H3 acetylation. Chromatin and transcriptional profiling indicate that development arrests prior to the mid-2-cell stage, mediated in part by reductions in beta-catenin and MYC. These findings indicate that post-transcriptional regulation of tankyrase serves as a ligand-independent developmental mechanism for post-translational -catenin activation and is required to complete EGA. These studies shed light on basic genetic processes that can be disrupted by exposure to environmental chemicals and could impact on human fertility.