Localization and phosphorylation of HP1 proteins during the cell cycle in mammalian cells

Mammalian heterochromatin proteins 1 (HP1α, HP1β, and HP1γ) are nonhistone proteins that interact in vitro with a set of proteins that play a role in chromatin silencing, transcription, and chromatin remodeling. Using antibodies specific for each HP1 isoform, we showed that they segregate in distinct nuclear domains of human HeLa cells. By contrast, in mouse 3T3 interphase cells, HP1α and HP1β are strictly colocalized. In mitotic HeLa cells, all of HP1α and a fraction of HP1β and HP1γ remain associated with chromosomes. Immunostaining of spread HeLa chromosomes showed that HP1α is mainly localized on centromeres as shown previously for HP1β, while HP1γ is distributed on discrete sites on the arms of chromosomes. Biochemical analysis showed that HP1α and HP1γ are phosphorylated throughout the cell cycle, although more extensively in mitosis than in interphase, while HP1β apparently remains unphosphorylated. Therefore, despite their extensive sequence conservation, mammalian HP1 isoforms differ widely in their nuclear localization, mitotic distribution and cell cycle-related phosphorylation. Thus, subtle differences in primary sequence and in posttranslational modifications may promote their targeting at different chromatin sites, generating pleiotropic effects.