Fig 1: AtEPL1 interacts directly with the catalytic and platform subunits of Arabidopsis NuA4 and supports nucleosomal H4 and H2A.Z acetylation in vitro.a Pulldown assay showing the interaction between the recombinant AtEPL1 and HAM paralog pairs expressed in bacteria. Pull-down and input are abbreviated P-D and INP, respectively. b Interactions between AtEPL1s and HAMs were analyzed by far western blotting. After SDS–PAGE and western blotting of free GST and GST-AtEPL1A/B, the upper membranes were sequentially incubated with His-HAM1/2 and anti-His antibodies, whereas the lower membranes were incubated with an anti-GST antibody. The lower bands in the upper panels most likely indicate the interaction of HAM1/2 with partially degraded AtEPL1B. c Purification of recombinant A. thaliana picNuA4 from bacteria (Coomassie staining). A 6×His tag fused to HAM1 was used for purification on cobalt affinity resin. The numbers in the brackets indicate the expected molecular weights of the recombinant picNuA4 subunits. d Western blot showing Flag-HAM1 normalized protein levels in purified fractions from bacteria producing At picNuA4 complexes with and without AtEPL1A. Both fractions retained the Flag-HAM1 signal after ING2-His purification, indicating that AtEPL1A is not required for HAM1 binding. e In vitro assay of the HAT activity of recombinant A. thaliana (At) picNuA4 toward short oligonucleosomes (SONs) and free core histones (CHs). Recombinant S. cerevisiae (Sc) picNuA4 was used as a positive control. At-His corresponds to the purified His-HAM1 complex, and At-Flag indicates two versions of the Flag-HAM1-purified complex (see Supplementary Fig. 8a), with and without AtEPL1A, as shown in d. Autoacetylation on AtEAF6 is highlighted with an asterisk. f Immunoprecipitation (IP) of Flag-conjugated Arabidopsis H2A.Z protein (Flag-HTA9) from chromatin extract resulted in the isolation of human nucleosomes containing Arabidopsis H2A.Z. IP (1) and IP (2) correspond to Flag peptide eluates 1 and 2, respectively. SONs were used as a control. g In vitro assay of the HAT activity of recombinant A. thaliana picNuA4 toward Arabidopsis H2A.Z histone (FLAG-HTA9)-containing oligonucleosomes. Three versions of At picNuA4 were used, similar to the procedure in f. Only complexes containing AtEPL1A efficiently acetylated nucleosomal HTA9. Sc picNuA4 was used as a control with no activity towards HTA9. h Pull-down assay showing the interaction between AtEPL1B and a fragment of AtEAF1B corresponding to AAs 430-1322. HAM1 was used for comparison. All proteins were expressed in bacteria as either GST or 6×His fusions. Source data are provided as a Source Data file.