Total kinetic analysis reveals how combinatorial methylation patterns are established on lysines 27 and 36 of histone H3

Yupeng Zheng, Steve M.M. Sweet, Relja Popovic, Eva Martinez-Garcia, Jeremiah D. Tipton, Paul M. Thomas, Jonathan D. Licht, Neil L. Kelleher*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

101 Scopus citations


We have developed a targeted method to quantify all combinations of methylation on an H3 peptide containing lysines 27 and 36 (H3K27-K36). By using stable isotopes that separately label the histone backbone and itsmethylations,we tracked the rates of methylation and demethylation in myeloma cells expressing high vs. low levels of the methyltransferase MMSET/WHSC1/NSD2. Following quantification of 99 labeled H3K27-K36 methylation states across time, a kinetic model converged to yield 44 effective rate constants qualifying each methylation and demethylation step as a function of the methylation state on the neighboring lysine. We call this approach MS-based measurement and modeling of histonemethylation kinetics (M4K). M4K revealed that, when dimethylation states are reached onH3K27 or H3K36, rates of further methylation on the other site are reduced as much as 100-fold. Overall, cells with high MMSET have as much as 33-fold increases in the effective rate constants for formation of H3K36 mono- and dimethylation. At H3K27, cells with high MMSET have elevated formation of K27me1, but even higher increases in the effective rate constants for its reversal by demethylation. These quantitative studies lay bare a bidirectional antagonism between H3K27 and H3K36 that controls the writing and erasing of these methylation marks. Additionally, the integrated kinetic model was used to correctly predict observed abundances of H3K27-K36 methylation states within 5% of that actually established in perturbed cells. Such predictive power for how histone methylations are established should have major value as this family of methyltransferases matures as drug targets.

Original languageEnglish (US)
Pages (from-to)13549-13554
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number34
StatePublished - Aug 21 2012


  • EZH2
  • Epigenetics
  • Histone code
  • Mass spectrometry
  • Multiple myeloma

ASJC Scopus subject areas

  • General


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