A molecular examination of mRNA localization and cell polarization

描述（由申请人提供）：导致多种人类疾病的一个因素是细胞极性缺陷。许多细胞类型使用mRNA定位作为建立极性的手段。目前，我们对mRNA如何定位于高等真核生物的理解存在根本性的空白。该提案的目标是通过确定oskar和FMR1 mRNA定位的机制来解决这一知识缺口。oskar mRNA定位于果蝇卵母细胞，FMR 1 mRNA定位于神经元。这些转录本是研究的理想候选者，因为尽管它们位于不同的组织中，但它们的定位利用了许多相同的因素。因此，一个共同的机制可能运作，以定位不同的mRNA。因此，从这些研究中获得的见解预计将普遍适用于我们的理解mRNA定位。我们建议使用果蝇模型系统进行我们的研究。该模型系统将使我们能够在整个生物体的背景下研究mRNA定位的过程。这项研究有三个主要目标。Aim 1的动力是我们最近发现，剪接体的核心蛋白（称为Sm蛋白）在剪接之外的mRNA定位中发挥作用。在本研究中，我们建议确定的机制，Sm蛋白定位奥斯卡mRNA的功能。在目的2中，我们提出确定奥斯卡和FMR 1 mRNA耦合到马达蛋白进行运输的机制。参与转运两种mRNA的主要马达是驱动蛋白重链（Khc）。然而，Khc不使用其典型的衔接子来结合oskar和FMR 1 mRNA。因此，目前尚不清楚Khc如何与这些转录本结合。我们推测，一个未知的适配器连接Khc oskar和FMR 1 mRNA。目标2的目标是识别这种未知的适配器。最后，目标3集中在我们意想不到的发现，一个单独的电机，细胞质动力蛋白，也在奥斯卡mRNA定位功能。动力蛋白在FMR 1 mRNA定位中的功能已经得到证实。在这个目标中，我们将精确地定义动力蛋白在运输oskar mRNA中的作用。此外，我们将测试的假设，一旦动力蛋白提供奥斯卡mRNA的后极，它执行一个额外的功能，在卵母细胞内维持不对称的内吞活性。这些目标的完成将显著推进我们对细胞如何构建具有定位能力的mRNP以及mRNP与马达蛋白偶联用于运输的机制的理解。最终，从这些研究中获得的知识将使我们能够更有效地治疗由mRNA定位缺陷和细胞极性引起的疾病。)

DESCRIPTION (provided by applicant): A contributing factor to a wide spectrum of human diseases is defective cell polarity. Many cell types use mRNA localization as a means to establish polarity. At present, there is a fundamental gap in our understanding of how mRNAs are localized in higher eukaryotes. The goal of this proposal is to address this gap in knowledge by determining the mechanism by which oskar and FMR1 mRNAs are localized. oskar mRNA is localized in Drosophila oocytes and FMR1 mRNA is localized in neurons. These transcripts are ideal candidates for study because although they are localized in different tissues, their localization utilizes many of the same factors. Thus, a common mechanism might operate to localize diverse mRNAs. Insights gained from these studies are therefore expected to be generally applicable to our understanding of mRNA localization. We propose to perform our studies using the Drosophila melanogaster model system. This model system will enable us to study the process of mRNA localization in the context of the whole organism. This study has three main objectives. The impetus for Aim 1 was our recent discovery that core proteins of the spliceosome known as Sm proteins have a role, outside of splicing, in mRNA localization. In the present study, we propose to determine the mechanism by which Sm proteins function in localizing oskar mRNA. In Aim 2, we propose to determine the mechanism by which oskar and FMR1 mRNA are coupled to motor proteins for transport. The primary motor implicated in transporting both mRNAs is Kinesin heavy chain (Khc). However, Khc does not use its canonical adaptor to bind to oskar and FMR1 mRNA. Thus, it is currently unknown how Khc binds to these transcripts. We hypothesize that an unknown adaptor links Khc to oskar and FMR1 mRNA. The goal of Aim 2 is to identify this unknown adaptor. Finally, Aim 3 centers on our unexpected finding that a separate motor, cytoplasmic Dynein, also functions in oskar mRNA localization. A function for Dynein in the localization of FMR1 mRNA has already been demonstrated. In this aim, we will precisely define the role of Dynein in transporting oskar mRNA. Furthermore, we will test the hypothesis that once Dynein delivers oskar mRNA to the posterior pole, it performs an additional function in maintaining asymmetric endocytic activity within the oocyte. Completion of these aims will significantly advance our understanding of how the cell builds a localization competent mRNP, and the mechanism by which that mRNP is coupled to motor proteins for transport. Ultimately, the knowledge gained from these studies will enable us to more effectively treat diseases that result from defective mRNA localization and cell polarity. )