In animal chromatin, a fraction of the histone 2A's and 2B's is covalently attached to the protein ubiquitin through an isopeptide linkage. The ubiquitin moieties of the H2A's and H2B's are found to be in rapid equilibrium with the pool of free ubiquitin, both in dividing cells such as L1210 and Chinese hamster ovary cells and in nondividing cells such as unstimulated lymphocytes. The synthesis of ubiquitin and the formation of ubiquitinated histones are not linked to DNA synthesis. All ubiquitinated histones, including the four H2A variants and the H2B's are absent from isolated metaphase chromosomes.