Fig. 4

JMJD6 is required for the association of SIRT1 with chromatin. a Immunoaffinity purification of JMJD6-containing protein complexes. Cellular extracts from 293T cells expressing FLAG (vector) or FLAG-JMJD6 were immunopurified with anti-FLAG affinity columns and eluted with FLAG peptides. The eluates were resolved by SDS-PAGE and silver-stained. The protein bands were retrieved and analyzed by mass spectrometry. b JMJD6 interacts with SIRT1 in U2OS cells. Immunoprecipitation assays were performed with antibodies against the indicated proteins, followed by immunoblot analysis in U2OS cells. c JMJD6 interacts with SIRT1 in A549 cells. d The molecular detail of the interaction between JMJD6 and SIRT1. Whole-cell lysates from U2OS cells transfected with empty vector, FLAG-JMJD6, FLAG-mutant, or FLAG-JMJD6-N expression constructs were prepared, and immunoprecipitation was performed with anti-FLAG, followed by immunoblot analysis with indicated antibodies. e JMJD6 is required for the recruitment of SIRT1 to chromatin. The nuclear-soluble fraction (N) and chromatin-bound proteins (P) of U2OS cells, transfected with control or JMJD6-specific siRNAs, were extracted, and western blot analysis was performed using antibodies against the indicated proteins