Enroth et al. BMC Cancer 2011, 11:450 http://www.biomedcentral.com/1471-2407/11/450 most relevant histone modifications, both in general and from a cancer perspective, are histone H3 lysine 4 tri- methylation (H3K4me3), which is found in promoters of active genes and histone H3 lysine 27 trimethylation (H3K27me3), which is preferentially associated with pro- moters of inactive genes [6]. SMYD3, a histone methyl transferase specific for H3K4, is over expressed in color- ectal, hepatic and breast cancers, suggesting that H3K4 hypermethylation can occur at promoters of oncogenes [7]. MLL, another H3K4 methyltransferase, is frequently translocated in various forms of leukaemia [8] and the polycomb protein EZH2, a H3K27 specific histone methyl-transferase, is altered in multiple types of cancer [9]. Furthermore, recent reports suggest that H3K27me3 mediated silencing of tumor suppressor genes is a fre- quent event in prostate cancer [10,11]. Human genome-wide binding profiles for H3K4me3, H3K27me3 and other histone modifications have been previously generated, but in most cases, cancer cell lines were used as biological material [10,12]. Although this offers important insights into the functionality of the histone marks, it might not be optimal when investigat- ing their importance in oncogenesis, as exemplified by the discrepancy between H3K27me3 profiles in normal tissue and cell lines of the same origin [13], or by increased DNA methylation in embryonic stem cells (ESCs) due to in vitro culture [14]. Therefore, in order to establish how chromatin is altered during the onco- genic process, histone modification profiles from normal and tumor tissue samples are most desirable. However, and to the best of our knowledge, this has only been reported in liver [15] and pheochromocytoma [16]. Colon adenocarcinomas display numerous epigenetic alterations, including hypermethylation of tumor suppres- sor genes and loss of imprinting at IGF2/H19 [17]. Inter- estingly, the major risk factor for CRC is age and epigenetic lesions accumulate with aging and contribute to cell transformation [18]. Despite the evident role of histone modifications in colon cancer, histone modification profiles have not been analyzed in coupled normal and tumor colon samples. Here we present the first genome-wide maps of H3K4me3 and H3K27me3 in paired normal and tumor samples from CRC patients. We identify previously unknown changes in the methylation statuses between the normal and tumor samples and correlate these to CRC- related pathway and function. Taken together, these data could be an important resource in understanding the epi- genetic alterations associated to CRC. Results H3K4me3 and H3K27me3 binding profiles in paired normal and tumor samples from CRC patients We generated enrichment profiles for H3K4me3 and H3K27me3 in tumor samples from two CRC patients, Page 2 of 13 for which the profiles in normal mucosa have previously been generated [13] and for the colon adenocarcinoma cell line HT29. After determining the enriched regions (Figure 1A) for each modification in normal and tumor samples (Table 1 and Additional file 1), they were anno- tated to human genes using the UCSC knownGenes databases [19] (Table 1) (Methods). Signal footprints were created around transcription start sites and both the H3K4me3 and H3K27me3 pat- terns were similar to those reported [20-22] in other cell types (Figure 1B). As a quality control, microarray expression data from public repositories for normal colon, CRC tumors and HT29 were used [23]. Although the expression data comes from unrelated patient mate- rial, the objective was to study genome wide trends in transcription given the epigenetic status and not single gene effects where expression from the same patient material is essential. The expression of H3K4me3 enriched genes were higher than average, while H3K27me3 genes tended to be silent (Figure 2A). Biva- lent and especially semi-bivalent (Methods) gene groups included fewer genes and were poorly expressed, although with higher variation in expression. The quality of our data is also supported by previous validation of H3K4me3 and H3K27me3 enriched regions by qPCR in normal colon [13] demonstrating that ChIP-chip analy- sis can be performed using human tissue as biological material. For regions with associated transcripts, gene ontology (GO) annotations were obtained and an analysis was performed for over/under representation of terms com- pared to a whole genome background (Figure 2B, Meth- ods). The H3K4me3 and H3K27me3 gene groups have opposite functions and bivalent and semi-bivalent gene groups lie closer in functional space to H3K27me3 than H3K4me3 genes, as also suggested by expression data. Therefore, bivalent and semi-bivalent genes were treated as part of H3K27me3 gene groups for most subsequent analysis. In general, H3K4me3 fractions displayed over- representation in metabolic processes while H3K27me3 fractions showed enrichment in developmental cate- gories. The overrepresented categories in H3K4me3 and H3K27me3 genes were very similar in the HT29 cell line. H3K4me3 and H3K27me3 profiles are similar between normal and tumor samples but distinct compared to CRC cell lines Using all probes we calculated the Pearson’s correlation between all combinations of histone modifications, patient material (normal/tumor) and cell line (Addi- tional file 2, Table S1). In all cases the correlation between any patient materials was always strongest when that material was compared to another patient