Fig. 5: PRMT3 methylates SLC25A1 at R282 and R285.
From: PRMT3-mediated post-translational adaptation to fasting regulates metabolic flexibility
a, b Western blot (a) and quantification (b) of SLC25A1 in eWAT of 12-week-old male B6N mice at ad libitum and fasted for 12 h with or without 30 mg/kg SGC707 (n = 3). AL, ad libitum; F-12, fasted 12 h. c Immunoprecipitation assays using Anti-FLAG antibody in 293 T cells transfected with FLAG-tagged SLC25A1 and HA-tagged PRMT3, and western blot analysis of HA and FLAG. d Immunoprecipitation assays using Anti-ADMA antibody and western blot analysis of FLAG in 293 T cells transfected with FLAG-tagged SLC25A1 and HA-tagged PRMT3, representative image from 3 independent repeats. e Immunoprecipitation assays using Anti-FLAG antibody in 293 T cells transfected with FLAG-tagged SLC25A1 and HA-tagged PRMT3 with or without SGC707 treatment, and western blot analysis of Anti-ADMA antibody, representative image from 3 independent repeats. f Protein sequence alignment of SLC25A1 from Drosophila melanogaster to Homo sapiens, indicating the conserved R282 and R285 sites. g Structural prediction of WT SLC25A1 (SLC25A1WT) and R282G/R285C (SLC25A1Mut) by AlphaFold 3. h Immunoprecipitation assays using Anti-FLAG antibody in 293 T cells transfected with PRMT3 and SLC25A1WT/SLC25A1Mut and WB analysis of ADMA confirmed that SLC25A1 mutation reduced its methylation, representative image from 3 independent repeats. (i) Immunoprecipitation assays using Anti-ADMA antibody in 293 T cells transfected with PRMT3 and SLC25A1WT/SLC25A1Mut and WB analysis of FLAG confirmed that SLC25A1 mutation reduced its methylation, representative image from 3 independent repeats. j Immunoprecipitation assays using Anti-FLAG antibody in 293 T cells transfected with PRMT3 and SLC25A1WT/SLC25A1Mut and WB analysis of MYC confirmed that SLC25A1 mutation reduced its interaction with PRMT3, a representative image from 3 independent repeats. (k, l) WB analysis (k) and quantification (l) of protein stability of SLC25A1WT and SLC25A1Mut with or without PRMT3 by CHX assay (n = 3). Data are mean ± s.e.m. Statistical significance was determined using an unpaired two-tailed Student’s t test. Source data are provided as a Source Data file.
