KDM3A catalyses the oxidation of acetyl-lysine to hydroxyacetyl-lysine on histone H3K9

Abstract Histone modifications, including N ε -lysine acetylation and methylation, play critical roles in the regulation of eukaryotic transcription. The addition of acetyl and methyl groups and removal of acetyl groups to histones involve redox-neutral reactions. Demethylation is O 2 -dependent, as reported for reactions catalysed by the 2-oxoglutarate-dependent hypoxia-inducible factor (HIF) hydroxylases, one of which is structurally related to the Jumonji-C (JmjC) histone demethylases. We screened for substrates of the HIF-regulated JmjC lysine demethylase KDM3A and unexpectedly observed that purified recombinant KDM3A catalyses oxidation of the N ε -acetyl group of the Lys-9 of histone H3 (H3K9ac) giving an N ε -hydroxyacetylated product (H3K9acOH). Here we show that N ε -hydroxyacetyl-lysine is recognized by proteins known to bind to H3K9ac, including histone deacetylases and the YEATS domain-containing AF9. Studies employing an N ε -hydroxyacetyl-lysine selective antibody and mass spectrometry support the cellular relevance of N ε -hydroxyacetyl-lysine. Our combined biochemical and cellular results provide evidence for an unanticipated O 2 -mediated link between histone lysine N ε -acetylation and JmjC catalysis.