Genome-Wide Chromatin Immunopreciptiation and Gene Expression Analysis Indicates That MMSET Is a Transcriptional Repressor in Vivo

Multiple myeloma (MM) is a malignancy of plasma cells characterized by frequent chromosomal translocations of the immunoglobulin heavy chain (IgH) locus. The multiple myeloma SET domain (MMSET) gene is a recurrent chromosomal partner in the t(4;14) translocation, and MMSET levels are elevated in these patients relative to other myeloma cases and normal cells. Previously, we showed that MMSET is a histone methyltransferase with specific activity for lysine 20 on histone H4 and acts as a transcriptional repressor when tethered to a model target gene. To reveal the function of MMSET in t(4;14) MM in vivo, we identified MMSET target genes in the KMS11 t(4;14) MM cell line. Chromatin from these cells was subjected to immunoprecipitation with a polyclonal anti-MMSET antibody in biological replicate, amplified by ligation-mediated PCR and hybridized to NimbleGen 2.7kB promoter arrays, which represent 24,659 human promoters. Data analysis using the MaxFour algorithm ranked putative binding sites based upon intensities of 4 consecutive probes. The top 2000 promoters identified from each experiment were combined to yield a list of 1,412 putative MMSET target genes. This list was analyzed using the DAVID program (david.abcc.ncifcrf.gov/). Genes bound by MMSET includes those implicated in antigen processing and presentation (p 1000). This supports the notion that MMSET represses target genes in vivo. Functional annotation of genes bound by MMSET and expressed at low levels showed over-representation of genes implicated in toll-like receptor signaling pathway (p 100 in the wild-type KMS11 cells, 35 genes were up-regulated and 20 genes were down-regulated (>1.5 fold) in the KMS11KO cell line. Among the 692 genes bound by MMSET and expressed at a level of ≤100, 9 genes were up-regulated in KMS11KO cells. The up-regulated genes (presumably bound and repressed by MMSET) were categorized in cytokine receptor (p