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区隔/定位是必不可少的。这一建议涉及到一个非膜性屏障，它将酵母受精卵的细胞质细分为不同亲本起源的区域。以前在动物细胞的细胞质中描述过纤维“隔离带”的例子，其中一些包括中间丝或肌动蛋白。我们在酿酒葡萄球菌中检测到并将研究的合子中间区复合体（ZMC）几乎是最适合分析的。ZMC包含分隔素，它让人想起有丝分裂细胞周期中位于母细胞和子细胞之间的芽颈处含有分隔素的结构。线粒体重新分配的检查点：正如所观察到的那样，初步数据导致假设ZMC延迟亲代线粒体的融合，从而确保线粒体基因组的单代遗传。为了验证这一假设，我们将1)了解ZMC中的septin是否具有所需的动态特性，2)确定septin突变体中线粒体基因组的单代遗传是否受到损害。多聚体重新分配的检查点：为了确定ZMC是否对多聚体重新分配的延迟负责，我们将询问它们重新分配的时间是否反映了1)它们的大小，2)隔层的功能，或3)细胞核（和纺锤极体）之间的接触，正如我们的初步数据所表明的那样。功能概论：为了进一步了解ZMC的重要性范围，我们将了解它是否控制细胞质中新颗粒的再分配，限制细胞皮层和核包膜膜蛋白的通量，以及在早期受精卵中观察到亲本液泡不相互交换成分。鉴于与芽颈的明显相似性以及初步观察，我们将确定ZMC是否划定了胞吐发生的皮质区。我们还将了解ZMC的片段是否经历了特征性的共价修饰，以及这些修饰是否对ZMC的组织和功能至关重要。提案的最后一部分涉及确定一种药物的靶标，氯非虫伦可以可逆地改变septin组合的结构并中断合子中线粒体的再分配。由于这种药物也干扰动物细胞的septin，鉴定其靶点将开辟与septin生物学相关的意想不到的研究途径。

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.