Identification of nanos mRNA localizatino factors in Drosophila

描述(申请人提供)：细胞内信使核糖核酸定位是在真核细胞中产生不对称性的一种保守机制。在果蝇卵母细胞的后极定位Nanos(Nos)基因及其翻译，对于胚胎前后轴的形成和功能生殖系的建立是至关重要的。在包含多个部分功能元件的nos 3‘非编码区中，后部定位由顺式作用信号引导。先前的研究表明，反式作用因子与这种定位信号结合，将nos RNA包装成核糖核蛋白(RNP)复合体，从而允许定位。然而，到目前为止，只有一个nos定位因子被鉴定出来，那就是果蝇异质核糖核蛋白(HnRNP)M同源物，Rumpelstiltkin(Rump)。此外，NOS的定位必须与翻译控制紧密结合，以确保Nos蛋白的后继表达。这是通过另一个包含卵巢和胚胎翻译抑制物结合位点的顺式作用序列实现的。这些抑制物共同阻止了非定位nos基因的表达。目前尚不清楚本地化因子和翻译抑制因子是如何协调调控nos基因的。这项工作的目标是确定RNA定位信号是如何被细胞定位机制的组件识别的，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种蛋白质Lost在NOS定位中的作用，并建立了Lost和Rump之间的遗传和生化相互作用。这些结果将促使进一步的研究，以确定Lost是否间接或直接调节nos的定位，并将评估Lost与Rump的相互作用以及潜在的翻译控制因素。消除丢失和残基并不能完全消除nos定位，这可能是由于nos定位信号中的功能冗余。这一结果激发了本建议的第二个目标，即识别与nos mRNA定位信号相互作用的额外因素，并有助于调节mRNA的定位和翻译。总之，这些实验将使我们能够确定mRNAs是如何包装的，以便在细胞内定位，以及RNA定位和翻译控制是如何协调调控的。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，这项拟议的工作将为正常过程中的缺陷如何导致癌症等疾病提供洞察力。

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.