ORGANIZATION OF CENTROMERIC DOMAINS IN HEPATOCYTE NUCLEI - REARRANGEMENT ASSOCIATED WITH DE-NOVO ACTIVATION OF THE VITELLOGENIN GENE FAMILYIN XENOPUS-LAEVIS

The existence of a function-dependent, nonrandom organization of chrom atin domains within interphase nuclei is supported by evidence which s uggests that specific chromatin domains undergo spatial rearrangement under conditions which alter gene expression. Exposure to estrogen of male Xenopus laevis hepatocytes in vitro results in de novo activation of vitellogenin mRNA production and vitellogenin protein synthesis an d provides an ideal model to study the association between chromatin o rganization and changes in gene expression. In a test of the hypothesi s that the de novo induction of vitellogenesis in male X. laevis is as sociated with a spatial rearrangement of specific chromatin domains, c entromeric regions were localized by immunofluorescent labeling of ass ociated kinetochore proteins in naive and in estrogen-treated, vitello genic cells. Analyses by confocal scanning laser microscopy of the thr ee-dimensional spatial distribution of kinetochores in estrogen-treate d male hepatocytes showed that a significantly greater proportion of s ignals was associated with the nuclear periphery than in non-estrogen- treated, naive male cells. In hepatocyte nuclei, quantification of kin etochore signal sizes using image analysis showed that these signals w ere fewer in number and showed greater variation in size than those of cells in metaphase, with larger signals exhibiting total normalized f luorescence intensities of two, three, four, and five times that assoc iated with kinetochore signals of metaphase cells. These observations are taken to reflect the existence of clustering of kinetochores and, by extension, of centromeres. in these cells. In summary, the results show that centromeric domains within interphase nuclei of Xenopus hepa tocytes occur as clusters and that these domains undergo spatial rearr angement under conditions which alter the transcriptional State of the cell. (C) 1995 Academic Press, Inc.