Serine 298 Phosphorylation in Linker 2 of UHRF1 Regulates Ligand-Binding Property of Its Tandem Tudor Domain

J Mol Biol. 2020 Jun 26;432(14):4061-4075. doi: 10.1016/j.jmb.2020.05.006. Epub 2020 May 16.


Ubiquitin-like with PHD and RING finger domains 1 (UHRF1) is an essential factor for the maintenance of mammalian DNA methylation and harbors several reader modules for recognizing epigenetic marks. The tandem Tudor domain (TTD) of UHRF1 has a peptide-binding groove that functions as a binding platform for intra- or intermolecular interactions. Besides the groove interacting with unphosphorylated linker 2 and spacer of UHRF1, it also interacts with di/tri-methylated histone H3 at Lys9 and DNA ligase 1 (LIG1) at Lys126. Here we focus on the phosphorylation of Ser298 in linker 2, which was implied to regulate the ligand-binding property of the TTD. Although the protein expression level of UHRF1 is unchanged throughout the cell cycle, Ser298 phosphorylated form of UHRF1 is notably increased in the G2/M phase, which is revealed by immunoprecipitation followed by Western blotting. Molecularly, while unphosphorylated linker 2 covers the peptide-binding groove to prevent access of other interactors, small-angle X-ray scattering, thermal stability assay and molecular dynamics simulation revealed that the phosphate group of Ser298 dissociates linker 2 from the peptide-binding groove of the TTD to permit the other interactors to access to the groove. Our data reveal a mechanism in which Ser298 phosphorylation in linker 2 triggers a change of the TTD's structure and may affect multiple functions of UHRF1 by facilitating associations with LIG1 at DNA replication sites and histone H3K9me2/me3 at heterochromatic regions.

Keywords: DNA methylation; isothermal titration calorimetry (ITC); molecular dynamics; phosphorylation; small-angle X-ray scattering (SAXS).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CCAAT-Enhancer-Binding Proteins / genetics*
  • DNA Methylation / genetics*
  • DNA Replication / genetics*
  • DNA, Intergenic / genetics
  • Epigenesis, Genetic / genetics
  • Histones / genetics
  • Humans
  • Ligands
  • Molecular Dynamics Simulation
  • Phosphorylation / genetics
  • Protein Binding / genetics
  • Scattering, Small Angle
  • Serine / genetics
  • Tudor Domain / genetics*
  • Ubiquitin-Protein Ligases / genetics*


  • CCAAT-Enhancer-Binding Proteins
  • DNA, Intergenic
  • Histones
  • Ligands
  • Serine
  • UHRF1 protein, human
  • Ubiquitin-Protein Ligases