SYNECHOCYSTIS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 单细胞蓝藻聚球藻。PCC6803是一种具有非常理想的分子遗传(例如，对基因组进行测序)的模式生物(Kaneko等人。1996年)，易于转化)、生理(例如，易于在纯培养中生长)和形态(例如，细胞直径约为1.5微米，因此适合定量三维(3D)超微结构分析)特征。这些特性使聚球藻成为一个理想的实验系统，可以解决我们长期存在的关于光合作用和类囊体膜的生物发生和组织的问题，类囊体膜是太阳能捕获和能量传递的场所。 关于集胞藻细胞中类囊体膜网络的生物发生和3D组织仍然是关键的悬而未决的问题。在过去的工作中，我们结合了用于超微结构研究的冷冻制备方法，包括高压冷冻和冷冻替代，与树脂包埋半厚(250 Nm)样品的电子断层扫描，连续薄层(60 Nm)重建分析，以及冷冻断裂场发射冷冻扫描电子显微镜，以更好地了解野生型的细胞质细节(van de Meen等人)。2006)和突变体(Mohamed等人2005)菌株。我们希望利用国家大分子成像中心的冷冻EM，包括断层扫描，对处于冷冻水合状态的完整野生细胞进行成像，以加强我们正在进行的研究。我们相信，如果成功，冷冻-EM和采集后分析的先进生物成像方法将有助于澄清关于类囊体膜组织及其与类囊体膜中心(即支持膜网络3D组织的细胞质结构)之间的关系的问题，并提高我们对类囊体膜系统与质膜相互作用的理解。有了这些数据，加上我们的遗传和生化方法，我们将处于更有利的地位，以确定膜生物发生的位置，并了解这些膜的3D秩序是如何在这个模式生物中保持的。这项工作的结果可以用来在更广泛的意义上揭示与原核膜生物发生有关的问题。

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. The unicellular cyanobacterium Synechocystis sp. PCC 6803 is a model organism that has highly desirable molecular genetic (e.g., the genome is sequenced (Kaneko et al. 1996), easily transformed), physiological (e.g., easily grown in pure culture), and morphological (e.g., cells are ~1.5 um in diameter and thus suitable for quantitative three-dimensional (3D) ultrastructural analysis) characteristics. These characteristics make Synechocystis an ideal experimental system to address our long term questions regarding photosynthesis and the biogenesis and organization of thylakoid membranes, the sites of solar energy capture and energy transduction. There remain key unanswered questions regarding the biogenesis and 3D organization of the thylakoid membrane network in Synechocystis cells. In past work, we combined cryo-preparation methods for ultrastructural investigations, including high-pressure freezing and freeze substitution, with electron tomography of resin embedded semi-thick (250 nm) samples, serial thin section (60 nm) reconstruction analysis, and freeze fracture field-emission cryo-scanning electron microscopy to better understand cytoplasmic details in wild type (van de Meen et al. 2006) and mutant (Mohamed et al. 2005) strains. We hope to make use of cryo-EM, including tomography, at the National Center Macromolecular Imaging to image whole, wild type cells in their frozen hydrated states in order to augment our ongoing research. We believe that, if successful, the advanced bioimaging methods of cryo-EM and post-acquisition analysis will help greatly in clearing up questions regarding thylakoid membrane organization and its associations with thylakoid centers (i.e., cytoplasmic structures that support the 3D organization of the membrane network) and improve our understanding of how the thylakoid membrane system interacts with the plasma membrane. With this data in hand, along with our genetic and biochemical approaches, we will be in a much stronger position to identify sites of membrane biogenesis and understand how the 3D order of these membranes is maintained in this model organism. Results from this work could be used in shedding light in a broader sense on questions related to prokaryotic membrane biogenesis.