Identification of nanos mRNA localizatino factors in Drosophila

描述（由申请人提供）：细胞内mRNA定位是真核细胞中产生不对称性的保守机制。果蝇卵母细胞后极的nanos（nos）mRNA的定位及其随后的翻译对于胚胎前后轴的形成和功能性生殖系的建立至关重要。后部定位由包含多个部分功能元件的nos 3'UTR中的顺式作用信号指导。以前的研究表明，反式作用因子结合到这个本地化信号包装成核糖核蛋白（RNP）复合物，允许本地化的nosRNA。然而，只有一个一氧化氮合酶的定位因子，果蝇异质核糖核蛋白（hnRNP）M同源物，Rumpelstiltskin（Rump），迄今已确定。此外，nos定位必须与翻译控制紧密耦合，以确保Nos蛋白的后续表达。这是通过另一个顺式作用序列来实现的，该顺式作用序列包含卵巢和胚胎翻译阻遏物的结合位点。这些阻遏物共同阻止了未定位的nos mRNA的表达。目前还不清楚定位因子和翻译抑制因子如何协同调控nosmRNA。这项工作的目标是确定mRNA定位信号是如何被细胞定位机制的组成部分识别的，以及RNA结合蛋白与这些信号的相互作用如何指导RNA定位。初步结果确定了第二个蛋白质，失去了，在NOS定位的作用，并建立失去和臀部之间的遗传和生化相互作用。这些结果提示进一步的调查，将确定是否失去的行为间接或直接调节NOS的本地化，并将评估失去与Rump和潜在的，翻译控制因素的相互作用。消除Lost和Rump并不完全消除nos定位，可能是由于nos定位信号中的功能冗余。这一结果激发了该提议的第二个目标，即确定与nos mRNA定位信号相互作用并有助于mRNA定位和翻译调节的其他因子。总之，这些实验将使我们能够确定mRNA是如何被包装用于细胞内定位的，以及RNA定位和翻译控制是如何协调调节的。控制胚胎发育的蛋白质的异常表达与许多疾病有关。通过了解细胞在发育过程中调节基因表达的机制，拟议的工作将提供有关正常过程缺陷如何导致癌症等疾病的见解。

DESCRIPTION (provided by applicant): Intracellular mRNA localization is a conserved mechanism for generating asymmetry in eukaryotic cells. Localization of nanos (nos) mRNA at the posterior pole of the Drosophila oocyte, and its subsequent translation, is critical for embryonic anterior-posterior axis formation and for establishment of a functional germline. Posterior localization is directed by a cis-acting signal in the nos 3'UTR comprising multiple, partially functional elements. Previous studies suggest that trans-acting factors bind to this localization signal to package nos RNA into a ribonucleoprotein (RNP) complex that permits localization. However, only one nos localization factor, the Drosophila heterogeneous ribonucleoprotein (hnRNP) M homolog, Rumpelstiltskin (Rump), has been identified thus far. Furthermore, nos localization must be tightly coupled with translational control to ensure posterior expression of Nos protein. This is achieved by another cis- acting sequence that contains binding sites for ovarian and embryonic translational repressers. Together, these repressers prevent expression of unlocalized nos mRNA. It is currently unknown how nos mRNA is coordinately regulated by localization factors and translational repressers. The goal of this work is to determine how mRNA localization signals are recognized by components of the cellular localization machinery and how the interaction of RNA-binding proteins with these signals direct RNA localization. Preliminary results identify a role for a second protein, Lost, in nos localization, and establish both genetic and biochemical interactions between Lost and Rump. These results prompt further investigations that will determine if Lost acts indirectly or directly to regulate nos localization, and will evaluate the interaction of Lost with Rump and potentially, translational control factors. Eliminating Lost and Rump does not completely abolish nos localization, likely due to functional redundancy in the nos localization signal. This result motivates the second aim of this proposal, to identify additional factors that interact with the nos mRNA localization signal and contribute to the regulation of mRNA localization and translation. Together, these experiments will allow us to determine how mRNAs are packaged for intracellular localization and how RNA localization and translational control are coordinately regulated. Abnormal expression of the proteins that control embryonic development is associated with a number of diseases. By understanding the mechanisms by which cells regulate gene expression during development, the proposed work will provide insight as to how defects in normal processes lead to diseases such as cancer.