.  What is Synechocystis sp. PCC 6803?

      Synechocystis sp. PCC 6803 has developed into a model cyanobacterium that scientists around the world are using. The strain was isolated from a fresh water lake, and was deposited in
the Pasteur Culture Collection (PCC) in 1968, and due to the work of Prof. Sergey Shestakov and coworkers in Moscow and Dr. John Williams (Michigan State University and Du Pont) in the early 1980s the strain was recognized to be spontaneuosly transformable, to integrate foreign DNA into its genome by homologous recombination (allowing targeted gene replacement), and to be able to survive and grow in a wide range of conditions. For example, Synechocystis sp. PCC 6803 can grow in the absence of photosynthesis if a suitable fixed-carbon source such as glucose is provided.

      Perhaps most significantly, Synechocystis sp. PCC 6803 was the first photosynthetic organism for which the entire genome sequence was determined. In 1996, Dr. Satoshi Tabata and coworkers at the Kazusa DNA Research Institute finished the genomic sequence of this organism and made the information available in very useful format on a website named CyanoBase (http://www.kazusa.or.jp/cyano/cyano.html). This information, coupled with the facile gene deletion strategy that is available for Synechocystis sp. PCC 6803, has made Synechocystis sp. PCC 6803 the cyanobacterium of choice for research in many groups. Deletion mutants lacking a specific gene have been created and analyzed for many different genes. For example, Synechocystis is used to study pigment (carotenoid and chlorophyll) synthesis and its regulation, tocopherol synthesis, carbon metabolism, respiration, photosynthesis and a host of other processes. The information gained not only is valuable for understanding the physiology of cyanobacteria, but also for that of other organisms such as plants.

.  What is the use of a complete genome sequence?

      First of all, a complete genome sequence can be used to predict the location of the genes. Synechocystis has about 3,260 genes that code for proteins. The protein sequence can be compared to known proteins from other organisms (the standard algorithm is BLAST; http://www.ncbi.nlm.nih.gov/BLAST, and the function of about half of the Synechocystis proteins can be predicted in this way. The other half (more than 1,000 proteins!) either resembles other proteins of unknown function, or does not closely resemble proteins in other organisms. The challenge is to determine the function of these unknown proteins, and this large challenge is gradually addressed by a number of different groups. [Figure 4] Annotation of the function of genes and their corresponding proteins is a scientific community effort, and freely accessible websites with information on a growing number of Synechocystis genes have been established. Examples include http://cyano.genome.ad.jp, http://www.kazusa.or.jp/cyano/Synechocystis/comments/index.html, and http://genolist.pasteur.fr/CyanoList. One of the main goals of this gene identification effort is to assemble a blueprint of the reactions that occur in the organism. Initial information on this assembly is available on metabolic reconstruction web sites, such as http://wit.mcs.anl.gov/WIT2, http://www.genome.ad.jp/kegg/metabolism.html, and http://ecocyc.org/ecocyc/metacyc.html. Such a blueprint not only is of enormous scientific importance, but also greatly facilitates many potential applications including metabolic engineering efforts, in which the organism is used to produce new or more valuable compounds.

.  Why a Genomes-to-Life Project (Microbial Cell Project) on
       Synechocystis?

      Based on the genomic sequence, it has become possible to put together a partial metabolic map of the reactions that are catalyzed by enzymes encoded in the genome. In this way, we have been able to put together a partial, qualititative scheme of reactions that occur in Synechocystis, focusing on reactions involving photosynthesis, respiration, and carbon fixation. The challenge now is to obtain quantitative values on the flow rate through different, intersecting reaction pathways. This work requires a combination of metabolite level analysis, electron flow measurements, and flux determinations, preferably of wild type in comparison with mutants that lack a specific step of a pathway. This is now being pursued.

      Another important functional parameter regarding regulation of cellular metabolism is the protein composition of the cell. Comparing relative amounts of specific proteins in wild type and mutants (or in one strain under different environmental conditions) provides clues about cellular responses to particular stimuli. Comparative proteome studies are fairly labor-intensive and time-consuming, but protein levels often are closely linked to the capacity of the corresponding enzymatic step, and therefore this information is functionally very relevant.

      To be able to properly interpret functional information, a thorough understanding of the structure and dynamics of the cell is critical. Even though cyanobacteria have been investigated structurally, a comprehensive investigation has been lacking. For example, even though the existence of an internal membrane system (thylakoids; photosynthetic complexes are found here) and of a cytoplasmic membrane has been long established in cyanobacteria [Figure 5, Figure 6, Figure 7], it is still unknown whether they originate from each other, and how membrane biogenesis proceeds. In this study, electron tomography is being used for a three-dimensional reconstruction of Synechocystis [Figure 8 (movie), Figure 9] and some of its mutants [Figure 10, Figure 11, Figure 12, Figure 13, Figure 14] will be undertaken, so that biogenesis and compartmentation issues can be addressed.