Proteins recognized by antibodies against isolated cytological heterochromatin from rat liver cells change their localization between cell species and between stages of mitosis (interphase vs metaphase)

Heterochromatin in the cell nucleus seems to concentrate various proteins, such as Drosophila heterochromatin protein 1, which maintain the repressed state of gene expression. However, it still remains obscure how protein com position related to chromatin structure is different between heterochromati n and euchromatin in interphase nuclei. We isolated cytological heterochrom atin from sonicated interphase nuclei obtained from rat liver cells and pre pared antisera against it. The dense heterochromatic bodies seen in the pre paration of intact nuclei were duplicated in a relatively pure form during the preparation of heterochromatin. In the sodium dodecyl sulfate-polyacryl amide gel electrophoresis (SDS-PAGE) analysis, differences between the frac tions of heterochromatin and euchromatin were noted by their protein compos ition. Isolated heterochromatin was then digested by DNase after partial di gestion with trypsin and its dense structure changed to become highly sensi tive to DNase, The prepared antibodies reacted with the heterochromatin reg ion of rat liver cell nuclei and isolated cytological heterochromatin; howe ver, they did not react with euchromatin. Using immunohistochemistry, the a ntibodies bound to each cell nucleus in all tissues observed; some cell typ es were distinguished by their differential stainability (e.g. staining in the cytoplasm). Staining of the mitotic cells showed that the proteins reco gnized by the antibodies were localized in the cytoplasm and, in part, on t he chromosomes. Based on the results of molecular cloning from rat liver cD NA library using the antibodies as a probe, it seemed that the antibodies m ainly recognized two proteins similar to arginase and general vesicular tra nsport factor p115, respectively. The results obtained from these experimen ts reveal that some proteins located in the heterochromatin of interphase l iver cell nuclei seem to play important roles in condensing a portion of th e chromatin structure during interphase and suggest that proteins composing heterochromatin might be changed according to cell types or the stage of t he cell cycle. (C) 1999 Harcourt Publishers Ltd.