Mass spectrometry (MS) based proteomics coupled with co-immunoprecipitation for Pi treated and TGF-β1 cells.

<p>(A) Protein complex from phosphate (3mM for 72 hours) and TGFβ (10ng/ml TGFβ1 for 4 days) treated cells were immunoprecipitated by DNMT1 antibody and resolved in SDS-PAGE gel which stained by Coomassie blue. Untreated cells were used as control. (B) A list of protein candidates identified through Mass spectrometry analysis showed the commonly enriched proteins through DNMT1 immunoprecipitation from Pi and TGF-β1 treated cells. (C-D) Confirmation of the interaction between DNMT1 and HDAC, the protein complex was immunoprecipitated by (C) HDAC2 antibody and detected by DNMT1 antibody or was immunoprecipitated by (D) DNMT1 antibody and detected by HDAC2 antibody. (E) qRT-PCR analysis showed the upregulated mRNA expression of HDAC2 in both Pi and TGF-β1 treated cells. Results were normalized to reference gene GAPDH (expression is presented as means ± s.d., n = 3 independent experiments, ***P<0.001).