Relocation of nucleolar proteins around chromosomes at mitosis. A study by confocal laser scanning microscopy

Abstract

The behaviour of nucleolar antigens known to associate with chromosomes at mitosis was investigated in mammalian cells (HeLa, HEp-2, PtK1, CHO) by immunofluorescence and confocal laser scanning microscopy. Serial optical sections through mitotic cells, from prophase to telophase, were used to generate three-dimensional images of the antigen distribution. Our results indicate that, at the onset of mitosis, these antigens leave the nucleoli in a highly ordered manner to form a network extending from the nucleoli towards the nuclear envelope. The migration begins at very early prophase, when the condensation of the chromosomes is not yet visible. After completion of the migration at late prophase, the labelling is found at the chromosome periphery. The antigens remain distributed as a sheath surrounding the chromosomes from prophase to telophase. Therefore, the proteins involved in the formation of this perichromosomal layer have different behaviour than those of the prenucleolar bodies. The antigens appear to interact strongly with chromosomes, since they are not lost during chromosome isolation in hypotonic buffer. Each chromosome is entirely covered from one telomere to the other, except in the centromeric region. Thus the relocation of these nucleolar proteins does not appear to be the result of a passive accumulation at the chromosome periphery, but seems rather to be due to an active targeting to specific sites. Consequently, these proteins may have a determining function in the progression of the cells through mitosis, possibly by participating in the protection and stabilization of the chromosomes.