HVEM TOMOGRAPHY OF BASAL BODIES IN MUTANT AND WILD TYPE CHLAMYDOMONAS

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 对基体和中心粒的组装知之甚少。 我们正在使用衣原体作为一个模型系统，以确定和表征分子的重要基础身体结构和功能。在过去的一年中，我们已经使用了改进的标本制备和双轴电子断层扫描的方法来研究的结构和组织的基体在单细胞的衣藻，衣原体reinhardtii。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中的新结构。断层扫描重建的δ微管蛋白缺失突变体，uni 3 -1的基体，已证实，基体主要含有双微管。此外，仅存在于野生型细胞过渡区的星状纤维在uni 3 -1基底体的核心内重复。该突变体的远端纹状纤维不完整，小根微管可能错位，并观察到多鞭毛细胞。 uni 3 -1的抑制子，命名为tua 2 -6，包含α-微管蛋白中的突变。结合δ-微管蛋白缺失tua 2 -6; uni 3 -1细胞构建两个鞭毛，但它们保留了基体结构和小根微管定位的缺陷。这些数据表明，特定微管蛋白亚型的存在直接影响衣原体基体和相关结构的组装和功能。这项工作已经发表（O 'Toole et al.，摩尔Biol. Cell 14：2999-3012，2003）。 我们现在正在对BALD 2基因和一个新基因TNS 1突变的菌株进行分析，TNS 1在基体组装中很重要。Dutcher实验室最近发现BALD 2基因编码ε-微管蛋白。为了进一步了解BALD 2在基体形态发生中的作用，我们正在分析具有新的bald 2等位基因以及部分恢复基体结构和功能的基因外抑制子的菌株。

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. 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.