Assembly and Regulation of Yeast Spindle Poles

酵母纺锤杆的组装和调节

基本信息

批准号：
9919582
负责人：
JENNIFER L GERTON
金额：
$ 32.38万
依托单位：
STOWERS INSTITUTE FOR MEDICAL RESEARCH
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-05-01 至 2022-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9919582
关键词：
Aneuploidy Binding Binding Proteins Biochemical Biochemical Genetics Body Size Cell Cycle Cell Nucleus Cell division Cells Centrosome Chromosome Pairing Chromosome Segregation Chromosomes Color Complex Congenital Abnormality Coupled Cues DNA DNA biosynthesis Data Defect Development Electron Microscopy Ensure Eukaryota Eukaryotic Cell Event Fission Yeast Fluorescence Resonance Energy Transfer Gene Dosage Genetic Genetic Materials Genetic Screening Genome Stability Goals Growth Human Image Imaging technology Lead Lighting Link Malignant Neoplasms Membrane Methods Microscopy Microtubule-Organizing Center Microtubules Mitosis Mitotic Modeling Modification Molecular Mutation Nuclear Nuclear Envelope Nuclear Pore Complex Organism Orthologous Gene Phenotype Phosphorylation Play Ploidies Process Proteins Proteomics Regulation Resolution Role Saccharomyces cerevisiae Saccharomycetales Seeds Structure System Testing Tissues Work Yeasts base cancer risk cell type daughter cell fungus gamma Tubulin genetic information human disease in vivo in vivo evaluation innovation insight mutant particle prevent protein complex protein protein interaction quantitative imaging spindle pole body transmission process

项目摘要

PROJECT SUMMARY Accurate transmission of genetic information is required for growth, proliferation and development of tissues and organisms. Faithful segregation of chromosomes involves many events, including the duplication of microtubule organizing centers (MTOCs), known as centrosomes in metazoans and spindle pole bodies (SPBs) in fungi, once and only once per cell cycle. In order for the cytoplasmic microtubule apparatus emanating from the MTOCs to access the chromosomes in the nucleus, the nuclear envelope (NE) must also be remodeled during cell division. In some cell types the NE entirely or partially disassembles while in others it remains intact. Centrosomes and SPBs adapt differently to these two mechanisms, as illustrated by the NE insertion of SPBs in both Saccharomyces cerevisiae and Schizosacchromyces pombe. Although centrosomes and SPBs are structurally distinct, both types of MTOCs duplicate, grow and interact with the NE. Defects in any of these events could lead to spindle errors, which often result in the gain or loss of a chromosome (aneuploidy). Aneuploidy in yeast often can be tolerated, but in humans it is frequently associated with cancer due to changes in uncovered recessive mutations or alterations in protein complexes. This proposal seeks to elucidate conserved principles used by the cell to restrict centrosome/SPB duplication to once per cell cycle, to ensure the MTOC reaches a size where nucleation capacity is sufficient for chromosome segregation and to insert or tether centrosomes/SPBs to the NE. Our innovative two-color structured illumination microscopy (SIM) with single-particle averaging (SPA) approach sets our work apart because we are able to resolve SPB features and duplication intermediates required for these events that were not observed using electron microscopy, biochemical, genetic or other super-resolution methods. In this work, we build upon the observations we have made and further extend imaging technology by pairing fluorescence resonance energy transfer (FRET) with SIM. This advancement allows us to study protein-protein interactions during SPB duplication and compare them to protein-protein interactions in a mature SPB to understand how centrosome formation is controlled. Our preliminary data suggests that physical interactions are dynamic and change throughout the SPB duplication cycle, an idea that we will further investigate by examining phosphorylation and other cell cycle-dependent modifications. Using SIM, we can visualize the SPB pore—the ring-like structure that anchors the soluble SPB in the NE. The mechanisms by which this structure and the related pore at nuclear pore complexes form is poorly understood. Because we can observe the SPB pore in yeast, we can dissect the molecular events used by cells to create this hole in the NE. Our overall objective is to determine mechanisms that coordinate centrosome duplication with DNA replication (Aim1), elucidate the molecular events that allow the SPB to insert into the NE (Aim 2) and study how SPB size and microtubule nucleation is controlled (Aim 3) using imaging in combination with genetic and molecular methods.

项目摘要生长，增殖和发展需要准确的遗传信息传播组织和组织。染色体的忠实隔离涉及许多事件，包括重复微管组织中心（MTOC），被称为中心体和主轴杆体的中心体（SPB）在真菌中，每个细胞周期一次和仅一次。为了进行细胞质微管设备从MTOC发出以进入核中的染色体，核包膜（NE）也必须在细胞分裂期间被重塑。在某些单元格中，NE完全或部分解散，而在其他单元格中保持完整。中心体和SPB的适应于这两种机制，如NE所示 SPB插入酿酒酵母和精神分裂症。虽然中心体 SPB在结构上是不同的，两种类型的MTOC重复，生长和与NE相互作用。缺陷这些事件中的任何一个都可能导致纺锤错误，这通常会导致染色体的增益或丢失（非整倍性）。酵母中的非整倍性通常可以耐受，但在人类中，它经常与癌症有关由于未发现的隐性突变或蛋白质复合物改变。该提议试图阐明单元用来将中心体/SPB复制限制为每个细胞周期一次的配置原理，确保MTOC达到核能足以足以进行染色体分离和向NE插入或束带中心体/SPB。我们创新的两色结构化照明显微镜（SIM）使用单粒子平均（SPA）方法可以使我们的工作与众不同，因为我们能够解决SPB 这些事件所需的功能和重复中间体未观察到使用电子设备显微镜，生化，遗传或其他超分辨率方法。在这项工作中，我们以我们进行了观察，并通过配对荧光共振能量进一步扩展了成像技术用SIM转移（FRET）。这种进步使我们能够研究SPB期间的蛋白质蛋白质相互作用重复并将它们与成熟SPB中的蛋白质 - 蛋白质相互作用进行比较，以了解中心体形成受到控制。我们的初步数据表明，物理互动是动态的，并且改变了在整个SPB重复周期中，我们将通过检查磷酸化和其他依赖细胞周期的修饰。使用SIM，我们可以可视化SPB孔 - 戒指状结构这将固定的SPB固定在NE中。该结构和相关孔的机制核孔复合物的形式鲜为人知。因为我们可以观察酵母中的SPB毛孔，所以我们可以解剖细胞在NE中创建该孔的分子事件。我们的总体目标是确定与DNA复制协调中心体重复的机制（AIM1），阐明了分子允许SPB插入NE（AIM 2）并研究SPB大小和微管成核的事件使用成像与遗传和分子方法结合的受控（AIM 3）。

项目成果

期刊论文数量（9）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Molecular model of fission yeast centrosome assembly determined by superresolution imaging.

DOI：
10.1083/jcb.201701041
发表时间：
2017-08-07
期刊：
The Journal of cell biology
影响因子：
0
作者：
Bestul AJ;Yu Z;Unruh JR;Jaspersen SL
通讯作者：
Jaspersen SL

The half-bridge component Kar1 promotes centrosome separation and duplication during budding yeast meiosis.

DOI：
10.1091/mbc.e18-03-0163
发表时间：
2018-08-01
期刊：
Molecular biology of the cell
影响因子：
3.3
作者：
Agarwal M;Jin H;McClain M;Fan J;Koch BA;Jaspersen SL;Yu HG
通讯作者：
Yu HG

The role of gene dosage in budding yeast centrosome scaling and spontaneous diploidization.

DOI：
10.1371/journal.pgen.1008911
发表时间：
2020-12
期刊：
PLoS genetics
影响因子：
4.5
作者：
Chen J;Xiong Z;Miller DE;Yu Z;McCroskey S;Bradford WD;Cavanaugh AM;Jaspersen SL
通讯作者：
Jaspersen SL

Redistribution of centrosomal proteins by centromeres and Polo kinase controls partial nuclear envelope breakdown in fission yeast.