PUI RESEARCH-MUW-BRANDON

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。ICSA是一种自动运输的外膜蛋白，定位于细菌的旧极；在宿主结肠上皮细胞中，肌动蛋白细丝在同一极组装，用于细胞内游泳和细胞间扩散。有几条证据表明，ICSA在分泌之前定位于细胞质膜的内面，而且ICSA在分泌之前必须定位于旧极。今年夏天，我们在一次筛查中利用这些发现来确定ICSA的假定极地目标。我们还对酵母中的四种蛋白质--Bud8p、Bud9p、Rax1p和Rax2p的靶向感兴趣，它们都参与了二倍体酵母细胞的两极萌发。Rax1p和Rax2p都靶向于二倍体细胞的两端，并且都参与了Bud8p和Bud9p对这些极点的靶向。有趣的是，Rax蛋白分别针对远端和近端的Bud8p和Bud9p(相对于最近的芽)。Rax1p和/或Rax2p的缺失导致芽蛋白的异常靶向，从而观察到出芽的轴向分布，而不是野生型二倍体细胞特有的振荡的双极分布。我们感兴趣的是Rax1p/Rax2p复合体的靶向分子机制，以及这些蛋白在Bud8p和Bud9p的差异靶向中所起的作用。2005年8月的近期目标是确定Rax1p和Rax2p在振荡、两极萌发中的功能。为此，我们正在对用于合成基因组阵列筛选的相应基因进行删除。随后，我们将应用SGA筛选Bud8p、Bud9p蛋白质来确定这些蛋白质在发芽过程中的功能，并应用酵母2-杂交筛选来确定Bud8p、Bud9p、Rax1p和Rax2p相互作用的蛋白质。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. IcsA, an autotransported outer membrane protein is targeted to the old pole of the bacterium; the same pole where actin filaments are assembled for intracellular swimming and intercellular dissemination in host colonic epithelial cells. Several lines of evidence suggests that IcsA is targeted to the inner face of the cytoplasmic membrane before secretion and that IcsA must be targeted to the old pole before it is secreted. During the summer we have exploited these findings in a screen to identify a putative polar target for IcsA. We are also interested in the targeting of four proteins in yeast--Bud8p, Bud9p, Rax1p and Rax2p that are all involved in bipolar budding in diploid yeast cells. Rax1p and Rax2p are both targeted to both ends of diploid cells and are both involved in the targeting of Bud8p and Bud9p to these poles. Interestingly, the Rax proteins target Bud8p and Bud9p to the distal and proximal poles (with respect to the most recent bud), respectively. Deletions in Rax1p and/ or Rax2p lead to aberrant targeting of the Bud proteins such that an axial distribution of budding is observed as opposed to the oscillating bipolar distribution of buds characteristic of wild type diploid cells. We are interested in the molecular mechanism for targeting of the Rax1p/Rax2p complex and the role that these proteins play in the differential targeting of Bud8p and Bud9p. The immediate goal for August, 2005 is to define the function of Rax1p and Rax2p in oscillating, bipolar budding. To this end we are making deletions in the corresponding genes for use in a Synthetic Genomic Array screen. Later, we will apply the SGA screen to the Bud8p, Bud9p proteins to determine the function of these proteins in budding and also apply the Yeast 2-Hybrid screen to define proteins with which Bud8p, Bud9p, Rax1p and Rax2p interact.