In situ visualization of rDNA arrangement and its relationship with subnucleolar structural regions in Allium sativum cell nucleolus

Abstract

We used a DNA-specific staining technique to show the two states of DNA component distributed in the nucleolar region of Allium sativum cells. One state is the extended DNA fiber, and the other is the condensed DNA clump. In situ hybridization demonstrated that the extended DNA fiber was an rRNA gene. Anti-fibrillarin antibody immunolabeling revealed that these rRNA genes were located in the dense fibrillar component near the fibrillar center, including at the periphery of the fibrillar center. None was in the dense fibrillar component far away from the fibrillar center. The condensed DNA clump was located in the fibrillar center. Further observations showed that the rRNA genes in the nucleolus were all arranged around the fibrillar center and associated with the DNA clumps in the fibrillar center. Results of statistical analysis showed that the distribution region of rRNA genes occupied about one-third of the total dense fibrillar component region. Ag-NOR protein showed a similar distribution pattern to that of rDNA. Immunolabeling of an anti-RNA/DNA hybrid antibody demonstrated that the transcription sites of rRNA were located at the periphery of the fibrillar center and in the dense fibrillar component near the fibrillar center, and these sites were consistent with the location and arrangement of rDNA shown in situ. These results demonstrated that transcription of rRNA takes place around the fibrillar center and at the periphery, whereas the dense fibrillar component that was far away from fibrillar center was the non-transcription region. The DNA clumps within the fibrillar center were probably the anchoring sites for rDNA arrangement.