Next-Generation Sequencing Analysis of DNA Methylation in Acute Myeloid Leukaemia

Abstract

DNA methylation has an emerging role in pathogenesis and diagnosis of acute myeloid leukaemia (AML). In order to obtain a global view of the DNA methylation changes in AML genome, we applied methylated DNA immunoprecipitation followed by next-generation sequencing (MeDIP-seq) to AML patients from 4 different cytogenetic subtypes [t(8;21), t(15;17), trisomy 8 and normal karyotype] and normal bone marrows (NBMs). The MeDIP-seq signals were quantified, generating DNA methylation profiles with a 100bp resolution. Whole-genome DNA methylation average was 3% less in AML than NBM. This limited global hypomethylation was observed mainly in SINE repeat sequences. DNA methylation of specific genomic regions e.g. promoters, gene bodies, CpG islands (CGIs) and CGI shores clearly discriminated AML from NBM and differentiated each AML subtype. The preferential methylation was found in CGIs located outside the promoters. Additionally, significant hypomethylated interspersed repeat sequences differentiated each AML subtype. The MeDIP-seq data was validated by bisulphite pyrosequencing and Illumina Infinium Array. Two genes (SPHKAP and DPP6) with significantly methylated promoters were also of interest and further analysis of their expression showed them to be repressed in AML. Transfection of cancer cell lines with vector expressing SPHKAP revealed 2 times increase in the intracellular level of sphingosine-1-phosphate as measured by mass spectrometry. Moreover, HHEX, which was one of the genes that showed differential intragenic methylation in trisomy 8 AML, was also differentially repressed in trisomy 8 AML. In conclusion, novel MeDIP-seq data was presented with CGIs methylation, intragenic DNA methylation and hypomethylated repeats were characteristic for each AML subtype.