Identification and localization of a nucleolin homologue in onion nucleoli.
A protein homologous to nucleolin, a major nucleolar protein with multifunctional features involved in pre-rRNA synthesis and early processing, has been identified and localized in situ in onion root meristematic cells by different techniques, which have included the use of an antibody raised against hamster nucleolin. The protein was identified on Western blots of nucleolar proteins as a 64-kDa band, by means of the anti-nucleolin antibody, bismuth staining, and the silver staining-nucleolar organizer (Ag-NOR) method. The experiments also suggested that nucleolin could be a target of these two cytochemical stainings. Although the 64-kDa band corresponds to a major nucleolar protein, it is a minor one among total nuclear proteins. The same techniques were used in situ at the ultrastructural level, and the immunogold detection of the nucleolin homologue was quantitatively evaluated. The protein accumulates in the transition area from nucleolar fibrillar centers to the dense fibrillar component, which is considered to be the structural result of ribosomal gene transcription. Out of this transition area, the dense fibrillar component may be divided into two regions, proximal and distal with respect to fibrillar centers, which show, respectively, the significant and unsignificant presence of nucleolin; we interpret this fact as the expression of the topological arrangement of pre-rRNA processing. Fibrillar centers themselves showed a weak but significant labeling with the anti-nucleolin antibody. However, bismuth staining was absent from the interior of fibrillar centers, indicating that the nucleolin in them is not phosphorylated. Ag-NOR staining uniformly covered fibrillar centers and the dense fibrillar component (at least in its proximal region), but it did not stain condensed chromatin inclusions in heterogeneous fibrillar centers, showing that the binding of nucleolin to chromatin is associated with its decondensation. This work provides additional evidence of the high phylogenetic conservation of molecular motifs which take part in ribosome biogenesis.
Martin
M
M
Centro de Investigaciones Biológicas (CSIC), Madrid, Spain.
Garcia-Fernandez
L F
LF
Díaz de la Espina
S M
SM
Noaillac-Depeyre
J
J
Gas
N
N
Javier Medina
F
F
D000490:Allium / Q000737:chemistry* / Q000648:ultrastructure; D001729:Bismuth; D015153:Blotting, Western; D002466:Cell Nucleolus / Q000737:chemistry*; D016253:Microscopy, Immunoelectron; D009687:Nuclear Proteins / Q000032:analysis*; D010750:Phosphoproteins / Q000032:analysis*; D016601:RNA-Binding Proteins; D016622:Silver Staining; D000097796:Nucleolin
D001729:Bismuth; D009687:Nuclear Proteins; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D000097796:Nucleolin