PMC

Red: hypermethylated in CMML samples; Green: hypomethylated in CMML samples.

β values are depicted using a pseudocolor scale (Red = Genes hypermethylated; Green = Unmethylated genes). Samples are color coded. The top bar beneath the dendrogram shows CMML and healthy donor samples. Second bar indicates CMML samples with or without TET2 mutation. Third bar indicates CMML samples with or without JAK2V617F mutation. The lower bar shows the β value of JAK2 CpG analyzed in the infinium array.

A) Hierarchical cluster analysis based on abnormally methylated genes between CMML samples with TET2 mutations and CMML samples without TET2 mutations. B values are depicted using a pseudocolor scale (Red = Genes hypermethylated; Green = Genes hypomethylated). Samples are color coded. The top bar beneath the dendrogram refers CMML or healthy donor samples, second bar indicates CMML samples with or without TET2 mutations and lower bar indicates cytogenetic risk of CMML patients. B) Box plots for hypermethylated genes in CMML samples with TET2 mutations (TET2-mut) with respect to CMML TET2 wild type (TET2-wt) samples. C) Box plots for hypermethylated genes in CMML TET2-wt samples respect to CMML samples with TET2-mut.

The percentage of 5 hmC, 5 mC and ratio between 5 hmC/5 mC were measured in 8 CMML TET2-mut and 5 TET2-wt patient samples using qPCR. Two CpG located 5′upstream (−38 and −244 bp respectively) to the CpG analyzed in the methylation array in the case of LAX1; one upstream CpG (59 bp) and another downstream CpG (58 bp) to the CpG in the case of SLC22A12 gene and one CpG downstream (5 bp) in the case of VHL gene were analyzed. Median values of percentage of 5 hmC, 5 mC or ratio between 5 hmC/5 mC are indicated and P values were obtained using the 2-tailed T test or U Mann Whitney test. CG: CpG dinucleotide included in the array; CCGG: CpG dinucleotide in which 5 hmC and 5 mC have been analysed and TSS: transcriptional start site.