A High-Resolution Landscape of Mutations in the BCL6 Super-Enhancer in Normal Human B-Cells

Abstract The super-enhancers (SE) of lineage-specific genes in B-cells are off-target sites of somatic hypermutation. However, the inability to detect sufficient numbers of mutations in normal human B-cells has precluded the generation of a high-resolution mutational landscape of SEs. Here, we captured and sequenced 12 B-cell SEs at single-nucleotide resolution from ten healthy individuals across diverse ethnicities. We detected a total of ∼9000 subclonal mutations (allele frequencies <0.1%); of these, ∼8000 are present in the BCL6 SE alone. Within the BCL6 SE, we identified three regions of clustered mutations where the mutation frequency is ∼7X10 -4 . Mutational spectra show a predominance of C>T/G>A and A>G/T>C substitutions, consistent with the activities of activation-induced-cytidine deaminase (AID) and the A-T mutator, DNA Polymerase η, respectively, in mutagenesis in normal B-cells. Analyses of mutational signatures further corroborate the participation of these factors in this process. Single base substitution signature SBS85, SBS37, and SBS39 were found in the BCL6 SE. While SBS85 is a denoted signature of AID in lymphoid cells, the etiologies of SBS37 and SBS39 are still unknown. Our analysis suggests the contribution of error-prone DNA polymerases to the latter signatures. The high-resolution mutation landscape has enabled accurate profiling of subclonal mutations in B-cell SEs in normal individuals. By virtue of the fact that subclonal SE mutations are clonally expanded in B-cell lymphomas, our studies also offer the potential for early detection of neoplastic alterations. Significance We used Duplex Sequencing to detect low-frequency mutations in the BCL6 super-enhancer locus in normal human B-cells. The landscape of pre-existing mutations is remarkably conserved across different ethnicities and reveals clustered mutational hotspots that correlate with reported sites of clonal mutations and translocation breakpoints in human B-cell lymphomas. This high-resolution genomic landscape revealed by Duplex Sequencing offers accurate and thorough profiling of low frequency, pre-existing mutations in normal individuals, and the potential for early detection of neoplastic alterations.