phosphorylated Heterochromatin protein HP1α, S97A mutant, full-length

The heterochromatin protein HP1α consists of an N-terminal disordered phosphorylation tail (N-tail), chromodomain (CD), central flexible hinge region (HR), and C-terminal chromo shadow domain (CSD). CD binds to the lysine9-trimethylated histone H3 (H3K9me3) tail in nucleosomes and CSD forms a dimer bridging two nucleosomes with H3K9me3. Ser97 in HR is phosphorylated in vitro by Casein Kinase (CK2). Since S97 phosphorylation is barely detectable in cells, we replaced Ser97 with Ala, showing that the mutant also forms LLPS at its high concentration as wild type. Therefore, the full length of HP1α_S97A was used as that of the wild type. HP1α has four successive serine residues in N-tail that are phosphorylated by CK2 in vivo. N-tail phosphorylation enhances the H3K9me3 binding and liquid-liquid phase separation (LLPS).