Fig. 5: Methylated Pontin binds FOXO3a through the arginine methyl-binding domain of FOXO3a.
From: Pontin arginine methylation by CARM1 is crucial for epigenetic regulation of autophagy
a Transcription factor enrichment analysis for the methylated Pontin-dependent genes using EnrichR software. b Binding motif analysis using methylated Pontin ChIP-seq data. Hypergeometric p-values were calculated. c Co-immunoprecipitation assay was performed to detect the interaction between Pontin and various autophagy-related transcription factors upon glucose starvation. d Co-immunoprecipitation assay was performed to detect the interaction between FOXO3a and Pontin in Pontin WT, RK or RA-expressing MEFs. e, f Co-immunoprecipitation assay of FOXO3a with Pontin in WT or Carm1−/− MEFs (e) or from nuclear extracts obtained from WT or Carm1−/− MEFs (f) following glucose starvation. g Co-immunoprecipitation assay was performed to detect the interaction between FOXO3a and Pontin in MEFs under glucose starvation after treating with CARM1-specific inhibitors, EZM2302 (100 nM) and EPZ025654 (100 nM) for 96 h. h Scheme of in vitro methylation assay for preparing non-methylated and methylated Pontin (above). GST-pulldown assay was performed to examine the binding of FOXO3a to methylated Pontin or non-methylated Pontin (below). i Immunodot blot assay for the binding of GST-FOXO3a to Pontin or methylated Pontin peptide. j Co-immunoprecipitation assay of Pontin with FOXO3a WT or aromatic mutants following glucose starvation. k GST-pulldown assay for the binding of methylated Pontin to FOXO3a C terminus (418–673 aa) or F640/F642L mutant. Source data are provided as a Source Data file.
