Role of Septins in Organelle Transfer

描述(由申请人提供)：在真核细胞中，大分子和细胞器的准确分隔/定位是必不可少的。这一建议涉及一种非膜屏障，它将酵母受精卵的细胞质细分为不同亲本起源的区域。纤维状“排斥区”的例子以前已经在动物细胞的细胞质中被描述过，其中一些包括中间丝或肌动蛋白。没有一个像我们在酿酒酵母中检测到并将进行研究的受精卵中区复合体(ZMC)一样易于分析。ZMC包括隔质，使人想起在有丝分裂细胞周期中位于母亲和女儿之间的芽颈上含有隔质的结构。线粒体重新分布的检查点：初步数据导致假设ZMC延迟了亲代线粒体的融合，从而确保了线粒体基因组的单亲遗传，正如已经观察到的那样。为了评估这一假设，我们将1)了解ZMC中的Septins是否具有所需的动态特性，以及2)确定Septin突变体中线粒体基因组的单亲遗传是否受到影响。多聚体重新分布的检查点：为了确定ZMC是否导致多聚体重新分布的延迟，我们将询问其重新分布的时间是否反映了1)它们的大小，2)隔膜的功能，或3)核(和纺锤体极体)之间的接触，正如我们的初步数据所表明的那样。功能概括性：为了进一步了解ZMC的重要范围，我们将了解它是否调控新颗粒在细胞质中的重新分布，限制细胞皮质和核膜的膜蛋白流量，以及是否负责观察到早期受精卵中亲本液泡不相互交换成分。考虑到与芽颈的明显相似性以及初步观察，我们将确定ZMC是否划定了发生胞吐作用的皮质区域。我们还将了解ZMC的Septin是否经历了特有的共价修饰，以及这些修饰是否对ZMC的组织和功能至关重要。该提案的最后部分涉及确定一种药物的靶标，该药物可可逆地改变Septin组合的结构，并干扰受精卵中线粒体的重新分布。由于这种药物也干扰动物细胞间隔蛋白，因此识别其靶点将开启与间隔蛋白生物学相关的意外研究途径。 公共卫生相关性：我们将调查的隔板提供了将真核细胞的细胞质区域细分的屏障的原型。我们的研究将有助于理解这些非常普遍的原理，有助于阐明与几种疾病有关的间隔蛋白的功能，澄清对发育细胞生物学和线粒体基因组遗传至关重要的问题，并启动对控制细胞器分布的新检查点的研究。

DESCRIPTION (provided by applicant): Accurate compartmentation/positioning of macromolecules and organelles is essential in eukaryotic cells. This proposal is concerned with a non-membranous barrier that subdivides the cytoplasm of yeast zygotes into regions of distinct parental origin. Examples of fibrous "zones- of-exclusion" have previously been described in the cytoplasm of animal cells, some of which include intermediate filaments or actin. None is nearly as amenable to analysis as the zygote midzone complex (ZMC) that we have detected and will investigate in S. cerevisiae. The ZMC includes septins and is reminiscent of septin-containing structures at the bud neck that lie between the mother and daughter during the mitotic cell cycle. A Checkpoint for Redistribution of Mitochondria: Preliminary data lead to the hypothesis that the ZMC delays fusion of parental mitochondria and therefore ensures uniparental inheritance of the mitochondrial genome, as has been observed. To evaluate this hypothesis, we will 1) Learn whether septins in the ZMC have the dynamic properties that are required, and 2) Determine whether uniparental inheritance of the mitochondrial genome is compromised in septin mutants. A Checkpoint for Redistribution of Polysomes: To determine whether the ZMC is responsible for the delay in redistribution of polysomes, we will inquire whether the timing of their redistribution reflects 1) Their size, 2) Functionality of septins, or 3) Contact between nuclei (and spindle pole bodies), as is suggested by our Preliminary Data. Generality of Function: To further understand the scope of importance of the ZMC we will learn whether it governs the redistribution of novel particles in the cytoplasm, limits flux of membrane proteins of the cell cortex and nuclear envelope, and is responsible for the observation that parental vacuoles do not exchange components with each other in early zygotes. Given the apparent similarity to the bud neck as well as preliminary observations, we will determine whether the ZMC delimits a cortical zone within which exocytosis occurs. We will also learn whether septins of the ZMC undergo characteristic covalent modifications and whether these modifications are essential for the organization and functions of the ZMC. The final part of the proposal concerns identification of the target of a drug, forchlorfenuron that reversibly alters the structures of septin assemblages and interrupts redistribution of mitochondria in zygotes. Since this drug also perturbs animal cell septins, identification of its target will open unanticipated avenues of investigation related to septin biology. PUBLIC HEALTH RELEVANCE: The partition which we will investigate provides a prototype of barriers that subcompartmentalize regions of the cytoplasm of eukaryotic cells. Our studies will help understand these very general principles, help elucidate functions of septins which are implicated in several diseases, clarify issues which are central to developmental cell biology and inheritance of the mitochondrial genome, and initiate the investigation of novel checkpoints that govern the distributions of organelles.