Centrosomal mRNAs in Asymmetric Cell Division

细胞命运在发育过程中由两种机制指定：细胞-细胞信号传导和图案分子的不对称遗传。在这两者中，不对称细胞分裂的理解要少得多。私家侦探提出了进一步探索一种新的机制，不对称细胞分裂中发现的蜗牛Ilyanassa obsoleta，其中mRNA的图案基因定位于中心体在有丝分裂间期，然后不对称地继承在随后的division. First目标是进行原位杂交筛选，以找到定位模式之间的约2500 Ilyanassa mRNA，通过各种标准选择。该实验将显示中心体定位的mRNA的比例和功能类别，并确定mRNA不对称分离的细胞分裂集合。第二个目标是破坏局部分子的功能，从而剖析胚胎发育中这一过程的要求。到目前为止观察到的定位模式强烈表明，中心体mRNA的遗传是重要的模式。 这些实验将证明这一机制的功能意义，增加对早期Ilyanassa胚胎中基本模式化事件的理解，并作为新开发的基因破坏方法的试验场。初步研究将集中在中试筛选中发现的Ilyanassa Nanos蛋白，后续工作将检查其他候选细胞命运指定基因。智力价值：这些实验的结果应该大大提高我们对不对称细胞分裂新机制的理解，特别是关于其在发育中的功能作用。这一现象处于几个领域的交叉点，为解决这一问题而提出的实验本质上是跨学科的。这项研究将使用来自发育遗传学、经典胚胎学、细胞生物学、比较生物学和基因组学的工具和概念。更广泛的影响：这项工作预计将对理解组织的发育产生影响，其中不对称细胞分裂决定细胞命运，如神经系统。它也可能影响农业上重要的软体动物如双壳类的发育研究。这项工作将导致在公共数据库中沉积大量新的Ilyanassa obsoleta cDNA序列，这将有助于那些使用这种动物进行寄生虫学，生态毒理学，化学生态学和幼虫生态学研究的人。这项工作还将提供机会，让本科生参与研究，这项工作的结果将用于P.I.的发育生物学课程。最后，本文提出的实验将用于改进和验证研究Ilyanassa基因功能的方法，这将从根本上增强该系统的功能。

Project SummaryCell fates are specified during development by two mechanisms: cell-cell signaling and asymmetric inheritance of patterning molecules. Of the two, asymmetric cell division is much less well understood. The P.I. proposes to further explore a novel mechanism of asymmetric cell division discovered in the embryo of the snail Ilyanassa obsoleta, where mRNAs for patterning genes are localized to centrosomes during mitotic interphase, then asymmetrically inherited during the subsequent division.The first objective is to carry out an in situ hybridization screen to find localization patterns among approximately 2500 Ilyanassa mRNAs, chosen by various criteria. This experiment will show the proportion and functional categories of mRNAs that are centrosomally localized, and determine the set of cell divisions where mRNAs are asymmetrically segregated. This screen will also identify candidate cell fate specification genes.The second objective is to disrupt the function of localized molecules and thus dissect the requirements for this process in embryonic development. The localization patterns observed so far strongly suggest that the inheritance of centrosomal mRNAs is important for patterning. These experiments will demonstrate the functional significance of this mechanism, increase understanding of the basic patterning events in the early Ilyanassa embryo, and serve as a proving ground for newly developed methods of gene disruption in this system. Initial studies will focus on the Ilyanassa Nanos protein discovered in the pilot screen, and subsequent work will examine other candidate cell fate specification genes.Intellectual merit: The results of these experiments should dramatically enhance our understanding of a novel mechanism of asymmetric cell division, especially with regard to its functional role in development. The phenomenon lies at the intersection of several fields and the experiments proposed to address it are intrinsically interdisciplinary. This research will use tools and concepts from developmental genetics, classical embryology, cell biology, comparative biology, and genomics.Broader impact: This work is expected to have implications for understanding the development of tissues where asymmetric cell division determines cell fate, like the nervous system. It may also impact studies of development in agriculturally important molluscs like bivalves. The work will result in the deposition of a large number of new cDNA sequences for Ilyanassa obsoleta in public databases, which should aid those who use this animal for studies in parasitology, ecotoxicology, chemical ecology and larval ecology. The work will also provide opportunities to involve undergraduates in research, and results from this work will be used in the P.I.'s Developmental Biology course. Finally, the experiments proposed here will serve to refine and validate methods for studying gene function in Ilyanassa, which will fundamentally enhance the power of this system.