HVEM TOMOGRAPHY OF BASAL BODIES IN MUTANT AND WILD TYPE CHLAMYDOMONAS

突变型和野生型衣藻基底体的 HVEM 断层扫描

• 批准号: 8362527
• 负责人: SUSAN K DUTCHER
• 金额: $ 1.06万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362527
• 关键词: Affect; Algae; Alleles; Biological Models; Cell physiology; Cells; Cellular Structures; Centrioles; Chlamydomonas; Chlamydomonas reinhardtii; Data; Defect; Distal; Fiber; Flagella; Funding; Genes; Grant; Methods; Microtubules; Morphogenesis; Mutation; National Center for Research Resources; Positioning Attribute; Preparation; Principal Investigator; Protein Isoforms; Publishing; Research; Research Infrastructure; Resources; Role; Source; Specimen; Structure; Tubulin; United States National Institutes of Health; Work; alpha Tubulin; cell type; cost; electron tomography; high voltage electron microscopy; improved; kinetosome; mutant; novel; reconstruction; tomography

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Little is known about the assembly of basal bodies and centrioles. We are using Chlamydomonas as a model system to identify and characterize the molecules important for basal body structure and function. Over the past year, we have used improved methods of specimen preparation and dual-axis electron tomography to study the structure and organization of basal bodies in the unicellular alga, Chlamydomonas reinhardtii. This work has revealed novel structures in wild-type and in strains with mutations affecting specific tubulin isoforms. Tomographic reconstructions of basal bodies from the delta-tubulin deletion mutant, uni3-1, have confirmed that basal bodies contain mostly doublet microtubules. Moreover, the stellate fibers that are present only in the transition zone of wild-type cells repeat within the core of uni3-1 basal bodies. The distal striated fiber is incomplete in this mutant, rootlet microtubules can be misplaced, and multiflagellate cells have been observed. A suppressor of uni3-1, designated tua2-6, contains a mutation in alpha-tubulin. In combination with the delta-tubulin deletion tua2-6; uni3-1 cells build both flagella, yet they retain defects in basal body structure and rootlet microtubule positioning. These data suggest that the presence of specific tubulin isoforms directly affects the assembly and function of basal bodies and associated structures in Chlamydomonas. This work has been published (O'Toole et al., Mol. Biol. Cell 14: 2999-3012, 2003). We are now pursuing analysis of strains that have mutations in the BALD2 gene and a new gene, TNS1 that is important in basal body assembly. The Dutcher lab has recently found that the BALD2 gene encodes epsilon-tubulin. To further understand the role of BALD2 in basal body morphogenesis, we are analyzing strains with new bald2 alleles as well as extragenic suppressors that partially restore basal body structure and function.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 关于基体和中心粒的组装知之甚少。 我们使用衣藻作为模型系统来识别和表征对基础身体结构和功能重要的分子。在过去的一年里，我们使用改进的样品制备方法和双轴电子断层扫描技术来研究单细胞藻莱茵衣藻基体的结构和组织。这项工作揭示了野生型和具有影响特定微管蛋白亚型的突变的菌株的新结构。对 δ-微管蛋白缺失突变体 uni3-1 的基体进行断层扫描重建，证实基体主要含有双联微管。此外，仅存在于野生型细胞过渡区的星状纤维在uni3-1基体的核心内重复。在该突变体中，远端横纹纤维不完整，根微管可能错位，并且已观察到多鞭毛细胞。 uni3-1 的抑制因子，称为 tua2-6，包含 α-微管蛋白突变。与δ-微管蛋白缺失tua2-6相结合； uni3-1 细胞构建双鞭毛，但它们保留基体结构和根微管定位的缺陷。这些数据表明，特定微管蛋白亚型的存在直接影响衣藻中基体和相关结构的组装和功能。这项工作已发表（O'Toole et al., Mol. Biol. Cell 14: 2999-3012, 2003）。 我们现在正在对 BALD2 基因和新基因 TNS1 发生突变的菌株进行分析，TNS1 对基础体组装很重要。 Dutcher实验室最近发现BALD2基因编码ε-微管蛋白。为了进一步了解 BALD2 在基体形态发生中的作用，我们正在分析具有新的 bald2 等位基因以及部分恢复基体结构和功能的外源抑制子的菌株。