Intranucleolar localization of human proliferating cell nucleolar antigen p120

The human proliferation-associated nucleolar antigen p120 was localized to substructures within HeLa cell nucleoli by immunofluorescence and immunoelectron microscopy of cells whose nucleoli were segregated by drug treatment or extracted with nucleases. By indirect immunofluorescence, protein p120 was localized diffusely throughout all interphase nucleoli. However, high resolution immunoelectron microscopy demonstrated that protein p120 staining delineated a network of 20-30-nm diameter beaded fibrils distributed throughout the nucleolus. This distribution was unique compared to that of the nucleolar proteins p145, RNA polymerase I, or B23 which were examined simultaneously. Drug-induced segregation of nucleoli by actinomycin D or dichlorobenzimidazole riboside, followed by immunoelectron microscopy, indicated that protein p120 was concentrated at the periphery of the granular region in segregated nucleoli. In situ nuclease digestion of cells with DNase I and/or RNase A did not release p120 from the nucleolus. Instead, p120 immunoreactivity was retained within phase-dense residual nucleoli. These results provide evidence that protein p120 is associated with, and in fact delineates, a network of fibrils which is retained in the nucleolar residue fraction of proliferating cells.