ORB GENE REGULATION OF TRANSLATION

ORB基因翻译调控

• 批准号: 8171471
• 负责人: Paul D Schedl
• 金额: $ 0.24万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8171471
• 关键词: Axon; Binding; Biochemical; Candidate Disease Gene; Cell Nucleus; Cells; Complex; Computer Retrieval of Information on Scientific Projects Database; Coupled; Dendrites; Deposition; Development; Drosophila genus; Elements; Embryo; Ensure; Eukaryota; Event; Family; Feedback; Funding; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genetic Translation; Goals; Grant; Growth; Homeostasis; Institution; Learning; Location; Masks; Mating Types; Memory; Messenger RNA; Microtubule-Associated Proteins; Movement; Mutation; Neurons; Nurses; Oocytes; Ovarian; Ovary; Play; Process; Protein Biosynthesis; Proteins; RNA-Binding Proteins; Research; Research Personnel; Resources; Role; Signal Transduction; Site; Somatic Cell; Source; Synaptic plasticity; Testing; Translating; Translational Activation; Translational Regulation; Translations; United States National Institutes of Health; Untranslated Regions; Yeasts; daughter cell; egg; fly; in vivo; member; mutant; prevent; segregation

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. mRNA localization plays a pivotal role in the establishment of polarity in the Drosophila egg and embryo. It typically begins with the transcription and packaging of the localized mRNAs in RNP complexes in nurse cell nuclei. These complexes 'mask' the mRNAs, preventing them from being translated while in transit, and 'mark' them for localization. The masked mRNAs are transported through the nurse cells and deposited in the oocyte. Depending upon their cis-acting localization elements, the mRNAs are then targeted to specific locations in the oocyte where they are anchored to the cytosketal network. Translational activation of the localized message provides a spatially restricted source of the protein product. Alternatively the mRNAs can be stored in a masked form until an appropriate signal, such as egg deposition, activates translation. This provides a mechanism for coordinating the localized synthesis of the protein with other developmental events. While the importance of mRNA localization (coupled with 'on site' translational regulation) was first documented in fly ovaries and early embryos, it is now clear that this regulatory mechanism is employed in many different contexts and occurs in virtually every eukaryote. For example, in yeast, the asymmetric segregation of ash-1 mRNA to the daughter cell provides a mechanism for ensuring that mating type switching does not occur in that cell. mRNA localization in somatic cells also occurs during the processes of movement, differentiation or growth. In neuronal cells, mRNAs encoding MAP2, a dendrite-specific microtubule-associated protein, and aCaMKII accumulate in the dendrites, but not in the axons. Localized mRNA translation has also been implicated in synaptic plasticity and learning and memory. In all of these cases, it is likely that the steps involved in mRNA localization and translational regulation are similar in broad outline to those described above in fly ovaries The proposed studies focus on the Drosophila orb gene which is one of the key components of the fly ovarian mRNA localization machinery. Orb is a founding member of the highly conserved CPEB family of RRM RNA binding proteins. In previous studies, we have shown that orb functions at the last step in the mRNA localization process, regulating the translation of the mRNA once it is on site. Orb binds to the 3? UTR of many localized mRNAs in vivo oskar (osk) Bicudal-D (Bic-D), and K(10) and is required to promote the translation of these mRNAs. In addition, Orb protein autoregulates its own expression by binding to the 3? UTR of localized orb mRNA and activating translation. This positive feedback loop is critical for the proper expression of Orb in the developing oocyte. The goal of our studies is to better understand how Orb autoregulates its own expression and how it controls the on site translation of mRNAs encoding factors critical for establishing oocyte/nurse cell identity and defining the polarity of the egg chamber and embryo. Part of the answer to these questions will come from identifying and characterizing other genes that are important for Orb autoregulation and/or function. For this purpose we propose to isolate proteins that are associated with Orb in vivo and identify them by Mass Spec analysis. To validate these biochemical studies we have devised a sensitized orb mutant background. The sensitized background will permit us to test mutations in candidate genes which encode proteins that are associated Orb in vivo for genetic interactions with Orb.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 mRNA 定位在果蝇卵和胚胎极性的建立中起着关键作用。 它通常始于护士细胞核中 RNP 复合物中局部 mRNA 的转录和包装。这些复合物“掩盖”了 mRNA，阻止它们在运输过程中被翻译，并“标记”它们进行定位。 被掩蔽的 mRNA 通过护士细胞运输并沉积在卵母细胞中。根据它们的顺式作用定位元件，mRNA 然后被靶向卵母细胞中的特定位置，在那里它们被锚定到细胞骨架网络上。 局部信息的翻译激活提供了蛋白质产物的空间受限来源。 或者，mRNA 可以以掩蔽形式储存，直到适当的信号（例如卵沉积）激活翻译。这提供了一种协调蛋白质局部合成与其他发育事件的机制。 虽然 mRNA 定位（与“现场”翻译调节相结合）的重要性首次在果蝇卵巢和早期胚胎中得到记录，但现在很清楚，这种调节机制在许多不同的环境中都得到应用，并且几乎在每个真核生物中都存在。例如，在酵母中，ash-1 mRNA 与子细胞的不对称分离提供了一种机制，确保该细胞中不会发生交配类型转换。 体细胞中的 mRNA 定位也发生在运动、分化或生长过程中。 在神经元细胞中，编码 MAP2（一种树突特异性微管相关蛋白）和 aCaMKII 的 mRNA 在树突中积累，但不在轴突中积累。 局部 mRNA 翻译也与突触可塑性以及学习和记忆有关。 在所有这些情况下，mRNA 定位和翻译调控所涉及的步骤很可能与上述果蝇卵巢中的步骤大致相似 拟议的研究重点是果蝇 orb 基因，它是果蝇卵巢 mRNA 定位机制的关键组成部分之一。 Orb 是高度保守的 RRM RNA 结合蛋白 CPEB 家族的创始成员。 在之前的研究中，我们已经表明，orb 在 mRNA 定位过程的最后一步发挥作用，一旦 mRNA 到达现场，就会调节 mRNA 的翻译。 Orb 与 3 绑定？体内 oskar (osk) Bicudal-D (Bic-D) 和 K(10) 中许多局部 mRNA 的 UTR 是促进这些 mRNA 翻译所必需的。 此外，Orb 蛋白通过与 3? 结合来自动调节其自身的表达。本地化 orb mRNA 的 UTR 和激活翻译。这种正反馈回路对于 Orb 在发育中的卵母细胞中正确表达至关重要。 我们研究的目标是更好地了解 Orb 如何自动调节自身表达，以及如何控制 mRNA 的现场翻译，这些编码因素对于建立卵母细胞/护士细胞身份和定义卵室和胚胎的极性至关重要。这些问题的部分答案将来自于识别和表征对 Orb 自动调节和/或功能很重要的其他基因。 为此，我们建议分离体内与 Orb 相关的蛋白质，并通过质谱分析对其进行鉴定。 为了验证这些生化研究，我们设计了致敏 orb 突变体背景。 敏化背景将使我们能够测试候选基因的突变，这些基因编码与 Orb 体内相关的蛋白质，以与 Orb 进行遗传相互作用。