＜Chromosome Segregation in Bacteria＞

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Chromosome must be precisely segregated into daughter cells for bacterial cells to proliferate. Although prokaryotic cells does not have “nuclei” that exist in Eukaryote, the chromosomal DNA, which is about 1000 times longer than cell length, is tightly condensed as “nucleoid” within the cell.
How such a tighltly packed chromosome can be segregated into daghter cells? We attempt to reveal the mechanism of this active chromomal segregation system in bacterial cell.

1. Centromere Like Region in Bacteria

For long years, Chromosome segregation in Bacteria had been thought to be a passive event that chromosomal DNA migrates toward daughter cells according to extension of the cell membrane to which the origins of chromosomes are anchored (Replicon Model of Jacob, F. et al., 1963). Recent study, however, revealed that specific region on chromosome of E. coli can dynamically migrate  toward cell poles independently of the extension of the cell membrane. On the contrary to the eukaryotic cells in which each step of replication and segregation of chromosomal DNA are apparently distinguishable, in bacteria, specific region of chromosomal DNA migrates toward cell poles and partitioned during replication. In this migration event, repliucation origin, oriC and terminal, ter shows different dynamics. Replication origin regions migrate to cell poles promptly after its replication and stay there till cell division. On the other hand, ter region stayed still at mid cell until just before cell division. Is there a centoromere like region at replication origin in former case? We revealed that the origin region migrates to cell poles via “migS” region on chromosome (Fig1, Yamaichi, Y. and Niki, H., 2004). Further more the chromosome DNA showed dynamic structural change (Fig. 2???). Now we are exploring and analyzing the proteins that form complex with the migS region.

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2. A moter protein that moves DNA

To migrate the replicated DNA toward cell poles as described above, there must exist driving force produced by motor protein or other factor. Recently, ATPase proteins, which are essential for stable partitioning of low copy number plasmids, are focused as the motor protein for DNA segregation.
 We have analyzed intracellular dynamics of one of such proteins, SopA, and revealed its function (Hatano et al., 2007). SopA protein is essential for the partitioning of sex determinant factor, F plasmid in E. coli, and putative motor protein generates driving force in plasmid partitioning for its ATPase activity. We investigated molecular function as motor protein by observing simultaneously both SopA labeled with fluorescent protein and partitioning of plasmid DNA. We have revealed that SopA formed a discrete focus nearby cell pole and a filamentous structure extended like helices throughout the cell (Fig. 4). The focus of SopA oscillated within the cell, and this oscillation is related to direction of the movement of plasmd (Fig. 5). The focus of SopA appeared oscillating between the “nucleoid borders” in a cell. The result indicates that SopA may recognize and cluster at the positions of nucleoid borders during oscillation. To test this possibility, we observed localization of SopA-YFP in anucleated cells (DNA less cells). Surprisingly, SopA-YFP showed discrete focus and oscillated in anucleated cells as observed in wild type cells. Furthermore, we observed helical structure of SopA-YFP in anucleated cells. These results indicate that SopA can assemble at the specific subcellular positions where overlap with nucleoid boader. MreB protein, that determines the cell shape of bacteria, also forms helical structure in a cell. MreB deletion mutation change cell morphology from rod to round shape. In such sphere cell, however, SopA normally functioned for plasmid partitioning (Hatano T. and Niki H., unpublished). And SopA formed filamentous structure and discrete focus that oscillated in the round cell. These results indicate that SopA form self assembled focus and filaments independently of nucleoid or cytoskeletal protein for cell shape. The filamentous structure of SopA may function as railway for migration of the partitioning plasmid (Fig. 6).

＜ Novel Genetic System for Chromosome Study ＞

Yeasts are powerful genetic tool to identify new elements or new phenotypes in a wide range of biology. However, due to its small cell size, there is a limitation in detailed cell biological analysis. Schizosaccharomyces japonicus is an alternative fission yeast. Although this haploid organism is poorly developed as a genetic tool, its nearly twice large nuclear size and fibrously condensed mitotic chromosomes compare to the other yeasts can be more suitable model system to understand mechanism of chromosome organization in vivo, such as mitotic chromosome segregation or interphase chromosome compartment. Our final aim is to discover new mechanisms that regulate chromosome cycle through the isolation of mutants with novel cytological phenotypes. We are currently collecting mutants showing disorganized chromosome behaviors. In parallel, we are generating chromosomal markers to ease our analysis. Thanks to Broad Institute, S. japonicus genomic sequences are fully available, and we have already tagged several important factors to visualize those components.

国立遺伝学研究所
系統生物研究センター 原核生物遺伝研究室（仁木研究室）

〒411-8540 静岡県三島市谷田1111-1 ＞ 連絡先詳細および地図はこちら

Microbial Genetics Laboratory (Niki Laboratory)
Genetics Strains Research Center National Institute of Genetics

1111-1 Yata Mishima Shizuoka Japan P.O. 411-8540 > Contact details and Maps