Tio M, Zavortink M, Yang X, Chia W
Institute of Molecular and Cell Biology, National University of Singapore Campus, Singapore 117609.
J Cell Sci. 1999 May;112 ( Pt 10):1541-51. doi: 10.1242/jcs.112.10.1541.
Cellular diversity in the Drosophila central nervous system is generated through a series of asymmetric cell divisions in which one progenitor produces two daughter cells with distinct fates. Asymmetric basal cortical localisation and segregation of the determinant Prospero during neuroblast cell divisions play a crucial role in effecting distinct cell fates for the progeny sibling neuroblast and ganglion mother cell. Similarly asymmetric localisation and segregation of the determinant Numb during ganglion mother cell divisions ensure that the progeny sibling neurons attain distinct fates. The most upstream component identified so far which acts to organise both neuroblast and ganglion mother cell asymmetric divisions is encoded by inscuteable. The Inscuteable protein is itself asymmetrically localised to the apical cell cortex and is required both for the basal localisation of the cell fate determinants during mitosis and for the orientation of the mitotic spindle along the apical/basal axis. Here we define the functional domains of Inscuteable. We show that aa252-578 appear sufficient to effect all aspects of its function, however, the precise requirements for its various functions differ. The region, aa288-497, is necessary and sufficient for apical cortical localisation and for mitotic spindle (re)orientation along the apical/basal axis. A larger region aa288-540 is necessary and sufficient for asymmetric Numb localisation and segregation; however, correct localisation of Miranda and Prospero requires additional sequences from aa540-578. The requirement for the resolution of distinct sibling neuronal fates appears to coincide with the region necessary and sufficient for Numb localisation (aa288-540). Our data suggest that apical localisation of the Inscuteable protein is a necessary prerequisite for all other aspects of its function. Finally, we show that although inscuteable RNA is normally apically localised, RNA localisation is not required for protein localisation or any aspects of inscuteable function.
果蝇中枢神经系统中的细胞多样性是通过一系列不对称细胞分裂产生的，其中一个祖细胞产生两个具有不同命运的子细胞。在神经母细胞分裂过程中，决定因子Prospero在基底皮质的不对称定位和分离，对于实现子代同胞神经母细胞和神经节母细胞不同的细胞命运起着关键作用。同样，在神经节母细胞分裂过程中，决定因子Numb的不对称定位和分离确保了子代同胞神经元获得不同的命运。目前已确定的、负责组织神经母细胞和神经节母细胞不对称分裂的最上游成分由无定向基因（inscuteable）编码。无定向蛋白本身不对称地定位于顶端细胞皮质，在有丝分裂期间细胞命运决定因子的基底定位以及有丝分裂纺锤体沿顶端/基底轴的定向过程中都是必需的。在此，我们定义了无定向蛋白的功能结构域。我们发现，氨基酸252 - 578似乎足以实现其所有功能，但它的各种功能的精确要求有所不同。氨基酸288 - 497区域对于顶端皮质定位以及有丝分裂纺锤体沿顶端/基底轴的（重新）定向是必需且充分的。更大的氨基酸288 - 5,40区域对于Numb的不对称定位和分离是必需且充分的；然而，Miranda和Prospero的正确定位需要氨基酸540 - 578的额外序列。实现不同同胞神经元命运分化的要求似乎与Numb定位所必需且充分的区域（氨基酸288 - 540）一致。我们的数据表明，无定向蛋白的顶端定位是其所有其他功能方面的必要前提。最后，我们表明，尽管无定向基因的RNA通常定位于顶端，但RNA定位对于蛋白质定位或无定向基因功能的任何方面都不是必需的。
