Biomedical subjects

Toshiro Kobori

Publications and source records attributed to Toshiro Kobori.

5 recordsLinked to original sources

PubMed2007

Chromatin dynamics of unfolding and refolding controlled by the nucleosome repeat length and the linker and core histones.

Chromatin is composed of genomic DNA and histones, forming a hierarchical architecture in the nucleus. The chromatin hierarchy is common among eukaryotes despite different intrinsic properties of the genome. To investigate an effect of the differences in genome organization, chromatin unfolding processes were comparatively analyzed using Schizosaccaromyces pombe, Saccharomyces cerevisiae, and chicken erythrocyte. NaCl titration showed dynamic changes of the chromatin. 400-1000 mM NaCl facilitated beads with approximately 115 nm in diameter in S. pombe chromatin. A similar transition was also observed in S. cerevisiae chromatin. This process did not involve core histone dissociation from the chromatin, and the persistence length after the transition was approximately 26 nm for S. pombe and approximately 28 nm for S. cerevisiae, indicating a salt-induced unfolding to "beads-on-a-string" fibers. Reduced salt concentration recovered the original structure, suggesting that electrostatic interaction would regulate this discrete folding-unfolding process. On the other hand, the linker histone was extracted from chicken chromatin at 400 mM NaCl, and AFM observed the "beads-on-a-string" fibers around a nucleus. Unlike yeast chromatin, therefore, this unfolding was irreversible because of linker histone dissociation. These results indicate that the chromatin unfolding and refolding depend on the presence and absence of the linker histone, and the length of the linker DNA.

Animals↗

PubMed2006

Comparative structural biology of the genome: nano-scale imaging of single nucleus from different kingdoms reveals the common physicochemical property of chromatin with a 40 nm structural unit.

Genome function is closely linked to the higher-order chromatin structures. To reveal a structural basis for the interphase chromatin organization, the 'on-substrate' lysis procedure was applied to nuclei isolated from human HeLa cells, chicken erythrocyte cells and yeast Schizosaccharomyces pombe, which possessed different intrinsic properties of the genomes such as histone composition and inter-nucleosomal distance. The isolated nuclei on a coverslip were successively treated with a detergent and a high-salt solution to extract the nuclear membrane and the nucleoplasm, and therefore, atomic force microscopy (AFM) visualized the structural changes in response to the lysis procedure. After the nucleoplasm was extracted, AFM clarified that chromatin fibers, approximately 40 nm in width, were partially released out of the nuclei and that the other chromatin still remaining in the nuclei was composed of granular structures with diameter of 80-100 nm. Thus, these results suggest that the approximately 40 nm fiber would be a stable structural unit and fold the 80-100 nm granules into a one-step higher unit. A common mechanism could be implied regardless of the intrinsic properties of the eukaryotic genomes.

Animals↗

PubMed2003

Hierarchical chromatin structure of Schizosaccharomyces pombe revealed by atomic force microscopy.

Many structural studies on higher eukaryotic chromatin have been carried out, but chromatin structure in fungi remains unclear. Schizosaccharomyces pombe has been used for investigations of chromosome function; however, the structural details of S. pombe chromatin have not been clarified owing to its small nucleus. We used atomic force microscopy for nano-scale imaging of chromatin isolated from S. pombe. Topographic images indicated that nuclear chromatin contained at least three hierarchical structures: large-scale chromatin fibers, spherical domains in the fibers, and nodules in the domains. The average diameters of the domain and the nodule were 363 +/- 85.2 nm and 46.2 +/- 9.30 nm. Each structure comprising the hierarchy was similar to higher eukaryotic chromatin thus far observed, despite definite differences in chromatin organization at the nucleosomal level. The presence of histone H1 suggested that there might be an alternative to compensate for histone H1 lacking in S. pombe.

Cell Cycle↗

PubMed2003

Atomic force microscopy study of chromosome surface structure changed by protein extraction.

We applied atomic force microscopy (AFM) to investigate the surface structure of barley chromosome in combination with a chemical treatment method. As a result, we have obtained high-resolution topographic images of granular structures with a diameter of ca. 50 nm on the surface of critical-point dried metaphase chromosomes. Treatment with 2M NaCl significantly modified the chromosome surface structure: surface roughness was increased and chromosome thickness was decreased. The NaCl treatment extracted two major proteins with molecular weights of 4000 and 20,000 Da. These proteins might be belonging to non-histone protein families that do not contain any aromatic amino acid. The results demonstrate the advantage of the combined method of high-resolution AFM imaging and chemical treatments for understanding nano-scale surface structures of the chromosome.

Chromosome Structures↗

PubMed2002

Imaging of chromosomes at nano-meter scale resolution using scanning near-field optical/atomic force microscopy.

Topographic and fluorescent images of whole barley chromosomes stained with YOYO-1 were observed simultaneously by scanning near-field optical/ atomic force microscopy (SNOM/AFM). The chromosome was relatively smooth and flat in the topographic images and no significant difference in height was present between regions of high fluorescent and low fluorescent intensity in the chromosomes. The telomeric region, labeled by fluorescence in situ hybridization (FISH) method, was also observed by SNOM/AFM at high resolution, and fluorescent signals of the telomeric region were clearly defined on the topographic image of chromatin fibers on the chromosome at the nano-meter scale level. Although the telomeric signals were usually visualized as a single fluorescent region at the end of sister chromatids by conventional light microscopy, they were observed separately as two fluorescent regions, less than 100-200 nm distance, using the SNOM/AFM. The SNOM/AFM offers great potential in identifying particular single gene location on chromosomes in the near future.

Benzoxazoles↗