Leucine methylation of protein phosphatase PP4C at C-terminal is critical for its cellular functions

Biochem Biophys Res Commun. 2014 Sep 12;452(1):42-7. doi: 10.1016/j.bbrc.2014.08.045. Epub 2014 Aug 15.


Background: Protein phosphatase 4 (PP4) has been known to have critical functions in DNA double strand break (DSB) repair and cell cycle by the regulation of phosphorylation of its target proteins, such as H2AX, RPA2, KAP-1, 53BP1. However, it is largely unknown how PP4 itself is regulated.

Methods: We examined the PP4C methylation on L307 at C-terminal by using methylated-leucine specific antibody. Then with PP4C L307A mutant, we explored that how nonmethylated form of PP4C affects its known cellular functions by immunoprecipitation, immunofluorescence, and DNA DSB repair assays.

Results: Here we show that PP4C is methylated on its C-terminal leucine residue in vivo and this methylation is important for cellular functions mediated by PP4. In the cells PP4C L307A mutant has significantly low activity of dephosphorylation against its known target proteins, and the loss of interaction between L307A PP4 mutant and regulatory subunits, R1, R2, or R3α/β causes the dissociation from its target proteins. Moreover, PP4C L307A mutant loses its role in both DSB repair pathways, HR (homologous recombination) and NHEJ (non-homologous end joining), which phenocopies PP4C depletion.

Conclusion: Our results demonstrate the key site of PP4C methylation and establish the physiological importance of this regulation.

Keywords: DNA damage response; Double-strand DNA breaks repair; Homologous recombination; Leucine methylation; Non-homologous end joining; Protein phosphatase 4.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • DNA Damage
  • DNA Repair
  • HeLa Cells
  • Humans
  • Leucine / metabolism*
  • Methylation
  • Phosphoprotein Phosphatases / chemistry
  • Phosphoprotein Phosphatases / genetics
  • Phosphoprotein Phosphatases / metabolism*
  • RNA, Small Interfering


  • RNA, Small Interfering
  • Phosphoprotein Phosphatases
  • protein phosphatase 4
  • Leucine