Genome architecture studied by nanoscale imaging: analyses among bacterial phyla and their implication to eukaryotic genome folding.

The proper function of the genome largely depends on the higher order architecture of the chromosome. Our previous application of nanotechnology to the questions regarding the structural basis for such macromolecular dynamics has shown that the higher order architecture of the Escherichia coli genome (nucleoid) is achieved via several steps of DNA folding (Kim et al., 2004). In this study, the hierarchy of genome organization was compared among E. coli, Staphylococcus aureus and Clostridium perfringens. A one-molecule-imaging technique, atomic force microscopy (AFM), was applied to the E. coli cells on a cover glass that were successively treated with a detergent, and demonstrated that the nucleoids consist of a fundamental fibrous structure with a diameter of 80 nm that was further dissected into a 40-nm fiber. An application of this on-substrate procedure to the S. aureus and the C. perfringens nucleoids revealed that they also possessed the 40- and 80-nm fibers that were sustainable in the mild detergent solution. The E. coli nucleoid dynamically changed its structure during cell growth; the 80-nm fibers releasable from the cell could be transformed into a tightly packed state depending upon the expression of Dps. However, the S. aureus and the C. perfringens nucleoids never underwent such tight compaction when they reached stationary phase. Bioinformatic analysis suggested that this was possibly due to the lack of a nucleoid protein, Dps, in both species. AFM analysis revealed that both the mitotic chromosome and the interphase chromatin of human cells were also composed of 80-nm fibers. Taking all together, we propose a structural model of the bacterial nucleoid in which a fundamental mechanism of chromosome packing is common in both prokaryotes and eukaryotes.

Overexpression of sigma factor, sigma(B), urges Staphylococcus aureus to thicken the cell wall and to resist beta-lactams.

Whole genome sequence analysis revealed that Staphylococcus aureus is provided with only a few sigma factors, including the alternative sigma factor, sigma(B), which is thought to regulate some stress responses. Since the sigB knock-out mutant did not show remarkable phenotypic difference, we constructed the over expressed mutant to examine the role of the sigB. Electron microscopic observation revealed that the mutant showed a variety of cell sizes compared with the parent strain which showed almost homogeneous cell sizes. The mutant delivered a thicker cell wall, about 20% thicker than the parent strain. It became resistant to the lytic activity of lysostaphin and also raised MICs to the cell-wall-affecting antibiotics. The yield of carotenoids and transcripts of pbps were also increased in the mutant. The result suggests that sigB plays some important roles in cell wall synthesis and in resistance to antibiotics that perturb the cell wall synthesis.

Studies on the constituents of Solidago virga-aurea L. III. Structures of solidagosaponins XXI-XXIX.

From the most polar fractions of Solidago virga-aurea L. (Compositae), 9 oleanane-type triterpene saponins named solidagosaponins XXI-XXIX (1-9) were isolated, together with a known saponin, virgaureasaponin 2 (10). These saponins are bisdesmosidic glycosides having two monosaccharides at C-3 and four or five monosaccharides at C-28. Their structures were established on the basis of spectroscopic and chemical evidence.