title: Three-dimensional architecture of ribosomal DNA within barley nucleoli revealed with electron microscopy.
abstract: To elucidate the topological positioning of ribosomal RNA genes (rDNA) and nucleolar structure in three dimensions, we examined the localization of rDNA using in situ hybridization (ISH) analysis by scanning electron microscopy (SEM). The rDNA genes within the three-dimensional architecture of nucleoli were detected on chromatin fibers that connect a thick strand-like structure and a protrusion of rDNA into the inner nuclear hole where the nucleolus is formed. This novel use of ISH together with SEM is useful for the analysis of nucleolar structure in detail. Furthermore, rDNA was detected at the periphery of the fibrillar centers (FCs) of the nucleolus using immuno-gold labeling together with transmission electron microscopy (TEM). In situ hybridization with TEM confirmed that rDNA is naked and thus active in the FCs of nucleoli; ISH with SEM confirmed that rDNA is not covered with ribonucleo proteins at the protruding point and is thus inactive. We also show that the distribution pattern of FCs differs from sample to sample. These results indicate that rDNA is transcribed dynamically in a time- and region-specific manner over the course of the cell cycle.
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Nara, Japan. m-iwano@bs.aist-nara.ac.jp
authors.forename: Megumi
authors.initials: M
authors.lastname: Iwano
authors.forename: Fang-Sik
authors.initials: FS
authors.lastname: Che
authors.forename: Seiji
authors.initials: S
authors.lastname: Takayama
authors.forename: Kiichi
authors.initials: K
authors.lastname: Fukui
authors.forename: Akira
authors.initials: A
authors.lastname: Isogai
mesh_terms: D002466:Cell Nucleolus / Q000648:ultrastructure*; D018744:DNA, Plant / Q000032:analysis; D004275:DNA, Ribosomal / Q000032:analysis*; D001467:Hordeum / Q000235:genetics* / Q000648:ultrastructure*; D017403:In Situ Hybridization; D008855:Microscopy, Electron, Scanning / Q000379:methods; D012335:RNA, Ribosomal / Q000235:genetics
chemical_list: D018744:DNA, Plant; D004275:DNA, Ribosomal; D012335:RNA, Ribosomal