Fig. 1: TG2 is a writer, eraser and exchanger of H3 monoaminylations.

a, TG2-mediated covalent modification of H3Q5 by 5-PT, and its visualization by CuAAC-meditated Cy5 conjugation. b, 5-PT pulse–chase in HeLa cells treated with or without TG2 inhibitors (ERW1041E or ZDON) after removal of 5-PT. Western blotting was performed for Cy5 (that is, H3 serotonylation) and TG2. c, 5-PT pulse–chase experiment in HEK293T cells transfected with WT TG2 and treated with or without TG2 inhibitors after removal of 5-PT. Western blotting was performed for Cy5 and TG2. d, Suggested mechanism for TG2-mediated H3 monoaminylation writing, erasing and exchange. e, LC–MS analysis of modified H3 peptides (as well as a deamidated H3Q5E peptide standard) (xi) after incubation with cellular lysates expressing WT TG2 versus TG2(C277A) (i–iv) or recombinant WT TG2 versus TG2(C277A) (v–x). H3Q5his peptide was incubated with WT TG2 in the presence or absence of replacement monoamine donors (xii (serotonin) versus xiii (dopamine)), demonstrating WT TG2-mediated deamidation of H3 monoaminylations in the absence of replacement donors (xiv), and exchange of H3 monoaminylations in the presence of replacement donors. Calculated (calc.) versus observed (obs.) masses are provided. High-performance LC (HPLC) UV traces, λ = 214 nm. f, WT TG2, but not TG2(C277A), transamidated histamine to H3Q5 on NCPs. NCPs premodified by histamine at H3Q5 could be deamidated by WT TG2, which was inhibited by treatment with ERW1041E. WT TG2 exchanged H3Q5his on NCPs in the presence of replacement donors, resulting in the establishment of H3Q5ser or H3Q5dop. NCP, nucleosome core particle. g, WT TG2, but not TG2(C277A), transamidated histamine to H3Q5 in HEK293T cells. H3Q5his-premodified histones could be deamidated by WT TG2 in cellulo, which was inhibited by treatment with ERW1041E. WT TG2 exchanged H3Q5his in cellulo in the presence of replacement donors, such as 5-PT (Cy5) or dopamine. H3 and actin were used as loading controls for western blotting. All of the experiments were repeated three times. Uncropped blots are shown in Supplementary Fig. 1.