New conserved candidate for egg activation and early embryogenesis in Drosophila.

描述（由申请人提供）：从卵到胚胎的过渡是发育的关键时期。由于这一过程对生育力至关重要，因此更好地了解其机制在发育生物学中很重要，也有助于优化ART程序。到目前为止，很难确定这种转变所需的关键蛋白质，部分原因是它太快而不涉及基因转录的变化。相反，许多证据表明，细胞状态的这种快速和全面的变化主要是通过现有蛋白质的翻译后修饰完成的。我们专注于蛋白质的磷酸化状态是在卵子激活过程中调制，推理这些蛋白质构成了一个丰富的池的重要分子的卵子到胚胎的过渡。我们建议使用生殖系特异性RNAi筛选258个保守的蛋白质，我们的蛋白质组学筛选发现在果蝇卵激活过程中磷酸化调节。通过利用该模式生物的最新菌株和方法进行直接的表型筛选，我们期望鉴定出调节从成熟卵母细胞到可以开始发育的“活化”卵的转变的基因，以及启动其胚胎发生的基因。我们将优先考虑筛选的顺序，优先考虑可能编码这种转变的上游调节因子的基因，以及一类下游效应因子：那些可能在细胞周期从减数分裂过渡到有丝分裂时调节细胞周期的基因。一旦我们确定了这些新的参与者，我们将使用蛋白质印迹和磷酸蛋白质组学来开始命令他们的行动。这项研究的目标是发现携带这种基因的关键基因。 从卵子到胚胎的重要发育转变。这些基因将允许未来的研究详细剖析进行卵子激活和启动胚胎发生的分子相互作用和途径。

DESCRIPTION (provided by applicant): The transition from egg to embryo is a critical time in development. Since this process is essential for fertility, a better understanding of its mechanisms is important in developmental biology and will also help to optimize ART procedures. It has thus far been very difficult to identify the critical proteins needed for this transition, in part because it is too rapid to involve changes in gene transcription. Instead, numerous lines of evidence argue that this rapid and comprehensive change in cell state is mostly accomplished by post-translational modification of existing proteins. We are focusing on proteins whose phosphorylation state is modulated during egg activation, reasoning that these proteins constitute an enriched pool of molecules important for the egg-to-embryo transition. We propose to use germline-specific RNAi to screen among 258 conserved proteins that our proteomics screens have found to be phospho-modulated during egg activation in Drosophila. By straightforward phenotypic screening that takes advantage of the latest strains and methods for this model organism, we expect to identify the genes that regulate the transition from a mature oocyte to an "activated" egg that can begin development, as well as those genes that initiate its embryogenesis. We will prioritize the order of screening, giving precedence to genes likely to encode upstream regulators of this transition, and to one class of downstream effectors: those likely to regulate the cell cycle as it transitions from meiosis to mitosis. Once we have identified these new players, we will use Western blotting and phosph-proteomics to begin to order their actions. The goal of the proposed research is to discover the critical genes that carry out the important developmental transition from egg to embryo. These genes will permit future studies to dissect in detail the molecular interactions and pathways that carry out egg activation and initiate embryogenesis.