This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor 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.
| Bucero, Marta Abril; Bajaj, Chandrajit; Mourrain, Bernard (2016) On the construction of general cubature formula by flat extensions. Linear Algebra Appl 502:104-125 |
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| Baker, Mariah R; Fan, Guizhen; Serysheva, Irina I (2015) Single-particle cryo-EM of the ryanodine receptor channel in an aqueous environment. Eur J Transl Myol 25:35-48 |
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