The relationship between Rab11-endosomes and the centrosome during division

摘要声明：纤毛病包括一组与遗传相关的疾病。 编码缺陷蛋白的突变，导致纤毛的异常形成或功能。 纤毛病与初级纤毛基底组织的分子通路缺陷有关 体/中心体和初级纤毛本身。患有纤毛病的患者可以表现出以下缺陷 左/右不对称、先天性心脏缺陷和多器官囊肿形成，包括 肝脏胰腺和肾脏在脊椎动物中，位于不对称器官中的活动纤毛起着 左右发展的重要作用。来自模式生物的证据，如斑马鱼的 不对称性（库普弗囊泡，KV），表明保守的纤毛驱动的左-右流动建立了左- 右信号调节靶基因以控制左/右不对称。该项目解决了 问题：纤毛细胞如何发育成功能性极化器官？我们建议这发生在 从细胞分裂、极性形成到纤毛形成 组装件.如果一个纤毛器官想要扩张它的中央腔，细胞需要重新进入细胞周期， 正确地将其有丝分裂纺锤体沿着平行于扩张管腔的最长轴定位。我们发现 Rab 11相关的内体和中心体蛋白cenexin都调节着 有丝分裂纺锤体和纤毛发生（Hehlynn和Doxsey，2014; Hung等人，2016年，但无论如何， 在这些过程中，没有被检查。在间期，Rab 11-内体 与中心蛋白所处的母体中心粒近远侧附属物相关联（Hehlynn等人，2012年）。 Rab 11和第二个GT3，Rab 8，可以直接与cenexin相互作用。此外，一个GT3级联 在纤毛发生和顶膜起始期间，Rab 11和Rab 8之间已经确定，但是如果 它需要中心体或发生在细胞周期的特定点是未知的。我们将测试 总体假设是Rab 11和/或Rab 8通过母体中心粒附属蛋白锚定， cenexin是纺锤体定位、极性形成和/或纤毛发生所必需的。为了验证这一点， 确定中心体是否可以调节Rab 11和/或Rab 8活性（Aim 1），并确定 当在细胞周期期间Rab 11和Rab 8活性需要调节组织形态发生时（Aim 2）。这将通过使用3-D肾上皮细胞培养模型和体内培养来实现。 发展模型，斑马鱼KV的形成，以确定一个框架，如何极性形成有关 对细胞周期的影响

SUMMARY STATEMENT: Ciliopathies comprise a group of disorders associated with genetic mutations encoding defective proteins, which result in either abnormal formation or function of cilia. Ciliopathies have been linked to defects in molecular pathways organized at the primary cilium's basal body/centrosome and the primary cilium itself. Patients suffering from ciliopathies can display defects in left/right asymmetry, congenital cardiac defects, and formation of cysts in multiple organs including liver, pancreas and kidney. In vertebrates, motile cilia located in an organ of asymmetry play an important role in left-right development. Evidence from model organisms, such as zebrafish organ of asymmetry (Kupffer's Vesicle, KV), indicates that conserved cilia-driven left-right flow establishes left- right signals to regulate target genes to control left/right asymmetry. This project addresses the question: How do ciliated cells develop into a functional polarized organ? We propose this occurs through a sequential process that starts with cell division, polarity formation, and finishes with cilia assembly. If a ciliated organ wants to expand its central lumen, cells need to re-enter the cell cycle and correctly position its mitotic spindle along the longest axis parallel to the expanding lumen. We found that both Rab11-associated endosomes and a centrosome protein, cenexin, regulate the positioning of the mitotic spindle and ciliogenesis (Hehnly and Doxsey, 2014; Hung et al., 2016), but whether they act together during these processes has not been examined. During interphase, Rab11-endosomes associate with mother centriole sub-distal appendages where cenexin resides (Hehnly et al., 2012). Rab11 and a second GTPase, Rab8, can directly interact with cenexin. In addition, a GTPase cascade has been identified between Rab11 and Rab8 during ciliogenesis and apical membrane initiation, but if it requires the centrosome or occurs during specific points of the cell cycle is unknown. We will test the overall hypothesis that anchoring Rab11 and/or Rab8 by the mother centriole appendage protein, cenexin, is required for spindle positioning, polarity formation and/or ciliogenesis. To test this we will determine whether the centrosome can modulate Rab11 and/or Rab8 activity (Aim 1) and determine when during the cell cycle Rab11 and Rab8 activity is required to regulate tissue morphogenesis (Aim 2). This will be accomplished by using a 3-D kidney epithelial cell culture model and an in vivo developmental model, zebrafish KV formation, to identify a framework for how polarity formation relates to the cell cycle.!