Genome-wide binding of phosphorylated MEL-18 protein at T334 in MDA-MB-231 cells

ChIP-seq analysis of MEL-18 WT- or MEL-18 T334A-overexpressing MDA-MB-231 cell lines. MEL-18, a core component of polycomb-repressive complex (PRC)-1, has been known to be phosphorylated at multiple residues in vitro; however, its functional roles in mammalian cells and human cancer remains largely unknown. Here, we examined the effect of MEL-18 phosphorylation at T334 site on polycomb-mediated epigenetic silencing in human breast cancer. Our results demonstrated that the phosphorylation of MEL-18 at T334 alters its genomic distribution and transcriptional activity that reflects functional change of MEL-18 in modulating breast tumour progression. Overall design: For the comparative analysis of the genomic binding patterns of MEL-18 wild type (MEL-18 WT) and its T334-phosphorylation-deficient mutant form (MEL-18 T334A), lysates from MDA-MB-231 cells stably expressing either FLAG-tagged MEL-18 WT or FLAG-tagged MEL-18 T334A were cross-linked, sonicated, and subjected to immunoprecipitation with anti-FLAG antibody. The ChIPed DNA and input DNA per each group were then sequenced using Hiseq2500 platform (Illumina)