DECIPHERING THE DNA DAMAGE RESPONSE IN THE GERM LINE BY PROTEOMICS

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 检测和消除DNA损伤的能力对于每个生物体的生存至关重要。因此，对DNA损伤的反应受到严格调控，需要几种信号转导途径的协调活性。许多参与相关信号传导过程的蛋白质已经在体外使用细胞培养物鉴定，并且众所周知体细胞中对DNA损伤的反应是如何调节的。但在这方面，生殖细胞系几乎没有受到影响，主要是因为它很难获得，而且材料数量非常有限。然而，最近的证据表明，体细胞和生殖细胞中的应激反应的调节之间存在重大差异。为了解开调节生殖细胞对DNA损伤的反应的信号网络，我们将应用质谱法并利用新兴的模式生物Nematostella vectensis。与哺乳动物相反，星状海葵Nematostella vectensis是为数不多的测序生物体之一，可以获得足够数量的卵母细胞的同质群体用于质谱分析。在这项研究中，我们将开发样品制备方法来富集信号蛋白，并使用MudPIT技术来鉴定蛋白质。由此产生的数据应使我们能够公正地看待体内遗传毒性应激后的细胞事件。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The ability to detect and eliminate DNA damage is essential for the survival of every organism. As such the response to DNA damage is tightly regulated and requires the coordinated activity of several signal transduction pathways. Many of the proteins involved in the involved signaling processes have been identified in vitro using cell cultures, and it is well known how the response to DNA damage is regulated in somatic cells. The germ line though has been left nearly untouched in this regard, mainly because of its difficult access and very limited amounts of material. Recent evidence however suggests major differences between the regulation of the stress response in somatic cells and that in germ cells. To unravel the signaling network regulating the germ line response to DNA damage, we will apply mass spectrometry and take advantage of the emerging model organism Nematostella vectensis. The starlet sea anemone Nematostella vectensis is in contrary to mammals one of the few sequenced organisms from which homogenous populations of oocytes can be obtained in amounts sufficient for mass spectrometry. In this study we will develop sample preparation methods to enrich for signaling proteins and use the MudPIT technology for identification of the proteins. The resulting data should allow us an unbiased look into the cellular events following genotoxic stress in vivo.