Control of efflux transporter activity by cell surface reorganization in embryos.

描述(申请人提供)：本研究的总体目标是了解早期胚胎发育过程中多种药物外排转运活动的细胞生物学调节。这一应用连接了两个知识体系，即主要在癌症和上皮运输背景下研究的多药外排转运的功能生理学，以及主要在细胞和发育生物学背景下研究的细胞表面和膜组织的结构变化。在这项应用中，候选人将描述在早期胚胎发育中发生的细胞表面变化与两种模式生物--海胆和小鼠--外排转运蛋白活性的伴随变化之间的关系。细胞生物学方面是要了解运输活动如何因细胞刺激而迅速改变，特别是结构(向微绒毛移动)和功能(防止毒物进入细胞)之间的关系。发育生物学方面是要了解未受精卵的膜是如何从专门与精子融合的膜变成现在保护和调节后续发育的膜。 这位候选人的博士后研究表明，在海胆卵受精后，ABCB(PGP)和ABCC(MRP)外排转运蛋白的活性大量而快速地上调。在大多数细胞中，多药转运体的插入是连续的，而在海胆中，这在受精后很快就会发生。因此，海胆为研究这一现象提供了一个强有力的模型。在前两个目标中，候选人将通过将转运蛋白移动到微绒毛顶端来表征受精后Sp-ABCB1a(哺乳动物PGP的同源基因)活性的重新分布(目标1)和Sp-ABCB1a在活胚胎中的定位动态(目标2)。此后，他将把这些发现扩展到小鼠模型，特别是跟踪小鼠卵母细胞受精后Pgp转运蛋白活性丧失的初步发现(目标3)。

DESCRIPTION (provided by applicant): The overall goal of this research is to understand the cell biological regulation of multidrug efflux transport activity during early embryo development. This application links two bodies of knowledge, the functional physiology of multidrug efflux transport, primarily studied in the context of cancer and epithelial transport, and the structural changes in cell surface and membrane organization, primarily studied in the context of cell and developmental biology. In this application, the candidate will characterize the relationship between cell surface changes that occur in early embryo development and the concomitant changes in the efflux transporter activity in two model organisms, the sea urchin and the mouse. The cell biology side is to understand how transport activity can be rapidly changed by cell stimuli and especially the relationship between structure (movement to microvilli) and function (prevention of entry of toxicants into the cell). The developmental biology side is to understand how the membrane of the unfertilized egg is altered from one specialized to fuse with the sperm to one that now protects and regulates subsequent development. The candidate's postdoctoral research has revealed massive and rapid up-regulation of ABCB (pgp) and ABCC (mrp) efflux transporter activity following fertilization of sea urchin eggs. In most cells insertion of multidrug transporters into the membrane is continuous, whereas in the sea urchin this occurs rapidly after fertilization. Thus, the sea urchin provides a powerful model for studying this phenomenon. In the first two aims, the candidate will characterize the post-fertilization redistribution of Sp-ABCB1a (an ortholog of mammalian pgp) activity by movement of the transporter to the tips of microvilli (Aim 1) and the dynamics of Sp-ABCB1a localization in living embryos (Aim 2). Thereafter, he will extend these findings to the mouse model, specifically following up on the preliminary finding of loss of pgp transporter activity after fertilization of mouse oocytes (Aim 3).